Exam 3 Flashcards
1
Q
The classification of viruses rest primarily on what 2 properties?
A
- Physical properties
- Chemical properties
**Physician is also required to know their disease potential and ways in which they can be controlled
2
Q
Definition of virus
- What is the classic definition of a virus? What does this mean?
- do they have both DNA and RNA? - What is the end goal of virus
- Examples of hosts that virus can replicate in? (6)
A
- OBLIGATE INTRACELLULAR PARASITE; they are ultra microscopic and possess only one type of nucleic acid (DNA or RNA)
**Obligate intracellular means it must get into the host cell to replicate (e.g bacteriophage - virus whether bacteria)
- End goal is to reproduce
- HUMANS, ANIMALS, insects, fungi, bacteriophage and plants
3
Q
Replication of virus
- Parent virus replicates in the cell to produce many progeny virus particles.
* Identify 2 infections that result in the production of progeny virus
**What 2 things combine to form progeny virus particles within an infected host cell
A
- LYTIC INFECTION; when virus enter cell, the cell bursts open (lyses)
- PRODUCTIVE INFECTION; produce large quantities of virus progeny). In the production of progeny virus, all the components of the parent virus (nucleic acid and proteins) must be synthesized in the new viral cell
**Production of VIRAL NUCLEIC ACIDS plus new VIRAL PROTEINS within an host cell combine to form progeny virus particles
4
Q
Structural features of extracellular virus
3 parts
- Protein shell. What forms this? 2 functions
- Composed of nucleic acid? (Be precise)
- Optional. Not all viruses have this. *what sensitivity does it account for? 2 functions? **what component on the envelope does all these?
A
- CAPSID; protein shell
- composed of subunits (Capsomers); many capsomeres = capsid.
- 2 functions; i) protect nucleic acid. II) important for attachment of virus to cell surface RECEPTORS - VIRAL GENOME; viral nucleic acid
- RNA or DNA but not both - ENVELOPE; optional membranous (lipid bilayer) outer structure of some viruses that surround capsid.
- accounts for ETHER SENSITIVITY of many enveloped viruses
- PROTEIN SPIKES on envelop are often involved in i) VIRUS ATTACHMENT to cell receptors and II) are IMPORTANT ANTIGENS
5
Q
Classification of virus into families
- What is classification based on? (Not based on?)
- Give examples of various classification of virus (9)
A
- Based on MORPHOLOGICAL and PHYSICAL (structure and biochemistry) features not the types of diseases that are produced
- Classifications
A. SIZE; poxvirus is largest, parvovirus is smallest
B. SHAPE of virus; icosahedral, complex, helical (envelop always)
C. NUMBER of capsomeres (adenovirus has 252 capsomeres)
D. TYPE of nucleic acid (DNA or RNA)
E. STRANDEDNESS; single or double
F. SIZE of nucleic acid (size affect viability)
G. SEGMENTATION of nucleic acid
H. Other criteria
I. Viral INFECTION of susceptible cells in tissue culture
6
Q
Classification of virus
- Size of virus
- which is largest
- which is smallest - Shape; 2 types
- which must always have envelope? Which s envelope optional?
- give 2 examples of complex shape - Numbers of capsomeres;
- which has 252 capsomeres? - Type of nucleic acid
- Examples of DNA vs RNA viruses - Strandedness
- which is mostly double stranded? Single stranded?
- exceptions?
A
- Size
- pox virus if largest
- parvovirus is smallest - Shape
A. Icosahedral; +/- envelop (most DNA virus)
B. Complex; Pox virus and retrovirus
C. Helical; must always have an envelope for helical animal virus (many RNA virus) - ADENOVIRUS has 252 capsomeres
- DNA or RNA nucleic acid but not both
- DNA; poxviridae > herpes viriridae > adenoviridae > papoviridae > parvo viridae
- RNA; rhamdoviridae, retroviridae, orthomyoviridae, paramyxoviridae, cornoviridae, picornaviridae, togaviridae - Most DNA is double stranded
- Most RNA is single stranded EXCEPT REOVIRUS which is double stranded RNA
7
Q
Classification of virus (cont’d)
- Size of nuclei acids
- the largest vs the smallest encode how many proteins each? - Segmentation of nucleic acid
- 8 segments vs single genome
A
- Poxvirus DNA is largest and encodes 300 proteins
- Parvovirus DNA is smallest and encodes 5 proteins
- *small virus = small DNA
- 1 - 10 segments, depends on particular virus. E.g
- influenza has 8 RNA segments
- parainfluenza has only a single genome
8
Q
Classification of virus (cont’d)
8. Other criteria - Genera, strains (7) A. What part of virus structure is sensitive to ether? Exception? B. Other susceptibility (4) C. 2 viral antigens D. Host cell specificity E. What does CPE show? F. Disease (same of different symptoms?) G. Genetic relatedness?
A
A. ETHER SENSITIVITY
- enveloped (Lipid bilayer) virus is usually sensitive except for POXVIRUS
B. Heat, chemical, pH sensitivity and detergent susceptibility
C. Antigenicity; 2 viral antigens are;
- capsid proteins
- envelop protein spikes
D. Host cell specificity
- cells in culture or cells within tissues in VIVO are susceptible to infection by certain viruses only; tropism
E. CPE - CYTOPATHIC EFFECT
- type of morphological changes of infected cells
- there are several types of CPE; lysis, slow cell death, enhanced cell proliferation
F. Disease in host organism (limited usefulness for classification, totally different viruses can cause diseases with same symptoms)
G. Genetic relatedness - hybridization, similarity of nucleotide sequences between the nucleic acids of 2 viruses
9
Q
Classification of virus cont’d
- Other criteria
A. what do you see in a confluence MONOLAYER - stained
B. confluence monolayer - not stained
C. Neutral red staining of confluence monolayer
D. Neutral red staining of single plaque
E. Neutral red staining of multiple plaques
A
A. what do you see in a confluence MONOLAYER - stained
- normal cells/tissues
- fibroblastic appearance of cells, elongated and spindle shaped
B. confluence monolayer - not stained
- see nothing
C. Neutral red staining of confluence monolayer
- viable/live cells absorb stain and acquire reddish appearance
D. Neutral red staining of single plaque
- PLAQUE FORMATION seen here; virus replicates in one cell - spread to surrounding cells (replicates) - kill so many cells over time - form PLAQUE (trace to first single cell)
- Point where they all meet (blue arrow) - PFU - plaque forming unit
E. Neutral red staining of multiple plaques
• This has multiple plaques
• Not all virus cause plaque formation. ONLY THOSE THAT CAN CAUSE LYTIC INFECTION OF CELLS
10
Q
Classification of virus cont’d
- Differentiate between constant inhibition and tumor derived cells
- other names
- growth or no growth?
A
- CONTACT INHIBITION (DENSITY DEPENDNET INHIBITON); contact each other and stop replicating
- Normal cells express contact inhibition - TUMOR/TRANSFORMED; cells contact each other but don’t stop replicating so grow in MULTILAYERS
- cells grow in multilayers or (loose contact inhibition and have altered growth characteristics)
11
Q
Classification of virus cont’d
- Viral infection of susceptible cells in tissue culture
A. Identify term - # of infectious virus particles added per cell to initial infection
B. What are 4 optimal conditions for infection
- what ions play role? What role?
- what inhibit attachment (4)
- what is term when virus required cells to be actively replicating
- term for cells that permit viral replication
A
A. MULTIPLICITY OF INFECTION (MOI); number of infectious virus particles added per cell to initiate infection
B. Optimal conditions for infections (4)
1. Ions; calcium and magnesium help virus attract to receptors on cells
- Inhibitors of adsorption (attachment); serum components, proteins, cross reacting antibodies; all use serumless media at time of infection
- Confluent vs non-confluent
- confluent; certain virus that require cells at specific state of division or metabolism
- non confluent; dont require actively diving cell - Permissive vs non permissive; influence lytic or productive infection
- permissive cells; support replication to produce progeny virus
- nonpermissive; lack essential factors for complete viral replication, may be transformed by certain tumor viruses
12
Q
CPE (cytopathic effects) are due to virus infection
**Identify the 3 types of CPE
A
- LYSIS (cytolytic); rapid destruction of cells
- termination of host cell protein and nucleic acid synthesis (e.g herpes) - Slow cell death; with continuous release of virus over a period of time (e.g measles)
- Enhanced cell proliferation; with or without virus production
- “transformed” cells; caused by DNA or RNA tumor virus - cells do not die as a result of viral infection (loss of contact inhibition lead to formation of multilayers)
13
Q
Identify the following infection types
- Small amount of virus produced but doesnt cause destruction of tissue around
- e.g chicken pox - varicella zoster (shingles caused by same virus that caused chicken pox when you are older)
A
- Persistent infection; Cell maintenance with or without obvious effect on the cell
- Latent infection; virus may no longer be evident yet viral genome is present
14
Q
Quantitation of viruses and detection of virus infected cells
- Virus assay for infectious virus particles
- 2 virus types
- PFU vs FFU (which is measure of infectious virus) - Virus assay for non infectious and infectious (no distinction)
- what shows how many viral particles
A
- Quantitation
A. CYTOLYTIC VIRUSES;
- can form plaques under agar (plaques are localized areas of dead cells in a cell monolayer)
- must be live cells because dead cells are not stained by neutral red
- a SINGLE infectious particle can initiate the formation of a single plaque
- one plaque = one infectious virus particle
***PFU (plaque forming units) - measure of infectious virus
1 PFU = 1 infectious virus
B. Virus with TUMOR POTENTIAL (can transform cells)
- FOCI; proliferation of transformed cels in localized areas of a cell monolayer
- FFU (focus forming unit) can be used to quantitate virus
- Virus assay - no distinction between infectious and non infectious
A. Hemagglutination; virus - erythrocytes lattice formation
- positive HA; diffuse coating of RBC on wells of plate
- negative HA; pallet/button at bottom of wells that indicate no viral-erythrocytes interaction and no lattice formation
- show how many viral particles
- Both dead and live virus cause hemagglutination. However not all viruses can cause hemagglutination
**some viruses have proteins on their surface that bind to receptors on specific RBC - this virus protein is called HEMAGGLUTININ
15
Q
Identify various methods of detecting infected cells
- RBCs attach to surface of infected cells vis viral proteins (hemagglutinin) expressed on cell surfaces
- • Instead of erythrocytes you add antibodies specific to viral protein with a fluorescent dye
• You are looking for fluorescence of infected cells - Amount of virus which kills 50% of animals
- 50% of cell culture show CPE
A
- HEMADSORPTION
- positive hemadsorption is when RBC attach to surface of infected cells by hemagglutin proteins - Immunofluorescence (If) and other staining procedures
- “If” requires fluorescent Ab which specifically binds to viral proteins on infected cells, irradiate fluorescein with UV light, gives off visible light
- cells stained with antibody-fluorescein look bright green under microscope - Death of animals (LD50); 1 LD50 equals the another of virus that kills 50% of animals
- Infection of cells in cultures, tissue culture infectious dose 50 (TCID50), 50% of cell cultures show CPE at a particular dilution amount of virus e.g 2 of 4 monolayer show CPE
16
Q
Viral Replication
Give an overview of the major events in the life cycle of infectious viruses (5)
A
- ATTACHMENT to receptors of susceptible cells
- PENETRATION and UNCOATING of nucleic acid (NA) - transport of nucleic acid to a specific site within the cell
- Intracellular SYNTHESIS; of virion components transcription - translation - lead to viral protein synthesis (translation) and viral NUCLEIC ACID SYNTHESIS
- ASSEMBLY/MATURATION; of virus components (viral proteins plus genomes at specific sites)
- RELEASE; of virions from the cell
17
Q
Virus replication (General Aspects)
- identify first step
A. where does it occur?
B. what bind to receptors on cell surface? (2)
C. what happens if cell surface lacks receptor?
A
- ATTACHMENT
A. Virus attach to receptors on outer surface of the cell
B. CAPSID and ENVELOP PROTEINS bind to the receptors on the surface of cells
C. Some cell types may lack receptors and therefore the virus will not attach and will not cause infection
18
Q
Virus replication (5 general aspects)
- Attachment
- identify/explain 6 factors that influence viral attachment to receptors - MOI
- Cell tropisim
- Ions
- No attachment inhibitors
- Temperature for attachment to occur
- Neutralizing antibodies
A
- MOI (multiplicity of infection); the number of virus infectious particles used to infect each cell
- the ratio of the quantity of virus particles : to infectious viruses (1:1 -> 2000:1) - Cell tropisim; ability to infect cell (cell must have receptor)
- Presence of appropriate receptors on cell surface - “specificity” between virus structural protein and cell receptor
- If there is no receptor - there will be no virus attachment to cell
* *Receptors determine tissue of CELL TROPISM - Ions (calcium and magnesium)
- proper ion concentration (Ca2+, Mg2+) helps binding of virus to receptors - No attachment inhibitors
- Lack of attachment inhibitors, remove serum proteins from media - Temperature for attachment to occur (4 to 37 degree Celsius)
- attachment occur over wide range of temp - Neutralizing antibodies (neutralize infectious activity)
- Neutralizing antibodies bind to surface proteins of the virus and prevent the virus from binding to the receptor so block attachment
19
Q
Viral replication (general aspects)
2a. Penetration
- Identify 2 ways in which virus penetrate into the cell
* *What is the end goal of virus
* *Degree range that penetration occurs
A
- Viropexis (phagocytic engulfement) or endocytosis
- Cell cytoplasmic membrane engulfs virus that is attached to receptor
- Virus is taken up in VACUOLES.
- The pH of endocytosis vacuole is important for UNCOATING - Fusion; second type of penetration
- pertains to ENVELOPED VIRUSES
- cell membrane fuses with viral envelope
- depend upon viral proteins which mediate fusion. Many fusion proteins must be proteolytically cleaved to cause fusion
**Penetration Occurs optimally at 37 degree and minimally or not at all at 4 degree.
** The end goal of the virus is to reproduce - it must introduce its nucleic acid inside the cell so it become dynamic and starts replicating
20
Q
Viral replication (general aspects)
2b. Uncoating
- describe process
**what 2 places in the cell can virus replication occur
A
UNCOATING; removal of some or all of capsid proteins - release genome in nucleus or cytoplasm (site of replication depends on virus under consideration)
- Some virus replicate in cytoplasm while some replicate in the nucleus
- Most DNA virus replicate in the nucleus (there is exception)
- Most RNA virus replicate in the cytoplasm
21
Q
Viral replication (general aspects)
- Synthesis
A. Transcription/translation of “parental” nucleic acid
B. Synthesis of viral nucleic acids (3 places for occurrence)
C. Transcription/Translation of “progeny” nucleic acid
- what is the order of steps?
- what are the products?
A
- PARENTAL
- occur before synthesis of viral nucleic acids
- transcription and translation of parental nucleic acid (viral genomes) to form EARLY mRNAs and EARLY proteins - Synthesis of viral nucleic acids (occurs in 3 options)
A. Cytoplasm (poliovirus)
B. Nucleus (herpesvirus)
C. Both sites (influenza virus) - PROGENY
- occur after synthesis of viral nucleic acid
- transcription and translation of progeny nucleic acid to form LATE mRNAs and LATE proteins
- translation occur in the cytoplasm but proteins can be transported to the nucleus or to the cell membrane depending on the details of replication of the virus under consideration
**The early and late designations relate primarily to DNA virus replication schemes
• Everything before nucleic acid synthesis is early
• Everything after nucleic acid synthesis is later
22
Q
Viral replication (general aspects)
- Maturation/Assembly
- what is the goal?
A. What come together to form progeny virus (2)
B. What is budding? What is the requirement to do budding?
- What is last step
A
- MATURATION/ASSEMBLY of viral components to form intact virions, occurs at specific sites within the cell
A. What come together to form progeny virus?
- Nucleic acid
- Capsid
B. Envelopment - “BUDDING” at cytoplasmic or nuclear membrane depending on virus
- pox virus is more complicated and has de novo synthesis of envelop
- only enveloped virus can do budding
- Virus is released or remain cell associated, depending on virus
23
Q
VIRUS PROGENY - different possibilities depending on virus family
Identify 3 possibilities
- location?
- 2 reasons you react non-infectious progeny
- 2 types of mutations (what is DI particles?)
A
- Extracellular or cell associated virus particles
- Infectious vs Non-infectious virus (2 reasons progeny is non-infectious)
A. Empty Capsids; no nucleic acid
B. Improper maturation; no proteolytic cleavage of envelop proteins - Mutants (2 types)
A. Drug resistance; mutation in viral nucleic acid yields i) viral proteins not affected by specific drugs II) nucleic acids not affected by a drug
B. DI particles (drug interfering particles); produced after high MOI of cells.
- DI particles are defective in that they lack portions of their nucleic acid (missing part of genome) - so can’t replicate on their own
- They require help form non defective “helper” non defective infectious virus
- DI particles interfere with replication of infectious virus; they compete for polymerase and may initiate persistent infections
24
Q
Identify the following concept
- Inclusion bodies (localize areas within infected cells cytoplasm or nucleus that contain viral particles)
- give examples - Granules and vacuoles - may or may not contain viral products
- Destruction or modification of normal cells structures (6)
A
Morphological and biological CHANGES OF INFECTED CELLS; CPE (cytopathic effect)
- Inclusion bodies
A. Negri - rabies (cytoplasmic)
B. Guarnieri - smallpox (cytoplasmic) - Granules and Vacuoles; may or may not contain viral products
- Destroy/modify structure of normal cell
A. Herpesvirus; breakdown of chromatin (cell DNA accumulate at periphery of nucleus)
B. Polio; dissociation of cell polysomes
C. cell and nuclear fusion syncytia; insert viral proteins into cytoplasmic/nuclear membranes corresponding to site of envelopment
D. increased methylation of tRNA, mRNA and rRNA
E. changing specificity of cell enzymes change energy generating systems
F. Cytoskeletal system (e.g microtubules) transform and dissociate
25
Identify 5 characteristics of mRNA
1. Use/function?
2. Capped at what end?
3. Polyadenylated at what end?
4. Outcome?
5. Possess what at 5’ end?
Characteristics of mRNA
1. Used to synthesize/translate proteins in cytoplasm
2. Capped at 5’ end
3. Polyadenylated at 3’ end
4. Outcome; may be synthesized as primary transcript - sliced to form mRNA - removal of introns and maintain Exons
5. May possess an UNTRANSLATED “LEADER’ sequence at 5’ end of mRNA
**There are atleast 6 mechanisms of transcription found among viruses (various ways viruses replicate)
• Some viruses (Ds DNA) has to transcribe (+) mRNA
• Some viruses (retrovirus - RNA) - reverse transcribe to DNA before transcribing to mRNA
• (-) RNA can’t serve as a messenger. It has to converted to the (+) to serve as messenger
26
Herpetoviridae - Herpesvirus family
Overview
A. structure?
B. Infect what type of cells? Type of infection
C. 4 specific members of the family (which is alpha, beta, gamma)
D. Which is a 5th member - important disease causing agent
E. Life threatening? Devastating to what people?
Herpesvirus family
A. members of family that cause disease in men have similar morphology (Ds DNA, Icosahedral capsid, envelop with glycoprotein spikes on its surface)
B. most infect cells of ectodermal origin and have capacity to establish latent infections (reactivated at later date)
C. **Herpes simplex (alpha), varicella zoster (alpha), cytomegalovirus (beta)
D. **Epstein-Barr virus (EB - gamma) is an important disease causing agent
E. These are non life threatening to most people but can be devastating to the immunocompromised
27
Herpes virus family (HSV - model system)
1. Replication of HSV
- location? (Site of viral replication)
- initial steps?
* *In HSV, What acts an a receptor? What is receptor called?
2. Viral DNA replication (3 aspects)
A. what happens to nucleus of linear genomic DNA?
B. How is concatemer formed?
C. Result?
1.
- HSV particle first attach to receptor on surface of permissive cell
- For HSV, HEPARAN SULFATE is believed to act as NECTIN 1 receptor
- After attachment, virus envelop fuse with cell membrane to enable penetration
- capsid containing the genomic DNA moves to the nucleus of the cell and genome is released in the nucleus
- **NUCLEUS is the SITE of VIRAL REPLICATION
2. Viral DNA replication
A. Inside the nucleus, the linear genomic DNA circularizes which results from terminal and internal nt sequence redundancies
B. Concatemer formation by ROLLING CIRCLE MECHANISM (similar to bacteriophage)
C. Specific cleavage of concatemer into unit size DNAs with reorientation, UL and Us, to yield the 4 configurations of DNA
28
Herpes virus family (HSV - model system)
3. Transcriptions and translation
- different classes of mRNAs appearing in a cascade fashion
A. Identify the 3 subclasses and state if they immerdiate early, early or late
B. Identify types of each subclasses
4. Assembly
A.
- alpha - immediate early
- beta - early
- gamma - late
* A temporal cascade of mRNAs and the corresponding proteins
B.
I) ALPHA PROTEINS are REGULATORY in nature
II) BETA PROTEINS are ENZYMATIC and are required for DNA replication (early proteins)
- **DNA dependent - DNA polymerase
- **Thymidine kinase
III) GAMMA PROTEINS; primarily STRUCTURAL and are used to form progeny virus, some are glycoproteins that are inserted into both the nuclear membrane and plasma membranes; by definition late proteins are synthesized following VIRAL DNA synthesis
4. Assembly
- capsid passes through nuclear membrane site. Acquires envelop from modified membranes of cytoplasmic vesicles having inserted of viral glycoproteins
29
Explains the reoccurrence of HSV (herpes simplex virus) in infection in individual that have a relatively high level of specific circulating serum antibody?
A. Ways in which virus can be reactivated
- Identify 3 stimuli and 1 infectious process?
B. How do you activate HSV from latency
C. Vascular stage and regression
D. Recurrent disease where?
1. Recurrent disease
a. In PRESENCE of Ab and cell immunity, virus can still be reactivated due to hormonal changes,
chemotherapy, mal-functioning immune system, aging, UV light, etc.
b.ACTIVATION of HSV from LATENCY: DNA harbored in
ganglia ---> activation ---> virus travel along
nerves back to original site of infection,
sequestered away from immune system
c.VESICULAR STAGE and REGRESSION, frequency of reactivation variable, may depend on the viral strain; eg. HSV-1 may exist as a variety of strains which differ in their pathogenicity
d. Recurrent disease in SKIN or EYE, OCULAR HERPES
infections can cause corneal blindness after numerous recurrences, cause stromal scarring
2. Immunity is not fully protective; recurrences still occu
A. Antibody important in resistance
B. Cellular immunity important in recovery
C. Vaccination, experimental and split vaccine needed; in subunit/split vaccine you strip off all the glycoproteins of virus
30
List 3 stimuli, including one infectious process, which are capable of inducing recurrent herpetic infections.
In PRESENCE of Ab and cell immunity, virus can still be reactivated due to
- hormonal changes,
- chemotherapy,
- mal-functioning immune system
- aging
- UV light, etc.
31
The following are primary disease characters of what viral infection
- gingivostomatitis, keratitis, skin, CNS affected
- transmitted through oral or genital secretions
HSV - Herpes simplex viruses
32
What is the significance of HSV type I and II in terms of disease producing characteristics
```
A. Location
B. Age of aquire
- gingivostomatitis
- eyes - dendritic keratitis
- skin - gladiatorum/whitlow/cold sore/edema herpeticum
- CNS ; neonate vs adult
- congenital
- Immunocompromised
```
**which condition is seen in both? (1.5)
General differences
A. HSV-1 (above waist)
- acquire HSV1 as a child ; most apparent 6-18 months
- Ab formation indicative of infection (can be asymptomatic)
- MOUTH; Gingivostomatitis, vesicular lesions in all parts of oral cavity
- EYES; dendritic keratitis; stromal involvement, HSV strain determined
- SKIN; Gladiatorum, herpetic whitlow, cold sores, eczema herpeticum
- CNS (*adult); spread along olfactory nerve - temporal lobe (neurogenic spread)
- IMMUNOCOMPROMISED; (AIDS patients) or malnourished - disseminated
B. HSV-2 (below waist)
- acquire AFTER PUBERTY through sexual contact and also at birth from infected birth canal
- CNS (*neonate); infected birth canal - transmission to neonate - viremia - brain
- CONGENITAL; fetus CNS, liver
- IMMUNOCOMPROMISED; (AIDS patients) or malnourished - disseminated
33
What considerations should be taken into account following discovery of genital herpes in the pregnant woman?
HSV-2 in pregnant woman
1. C section
- prevent possible fatal neo-natal infection (affect brain and visceral organs)
2. Acyclovir in 3rd trimester?
34
1. Sites for HSV to remain latent btw acute, clinically apparent episodes?
2. What anatomic/histologic xteristics
- at day 0-7
- after day 14
1. After primary infection HSV is transported up nerves to GANGLIA
A. Trigeminal ganglia (HSV1)
B. Sacral ganglia (HSV2)
2. Xteristics
A. At 0-7 day, VIRUS PRESENT in ganglionic cells as seen by ELECTRON MICROSCOPY
B. After 14 days; only HSV DNA present in cells - virus is not observed - episomal state of DNA in nuclei
35
Immunity and Epidemiology
1. Many adults with HSV -1 detected by what?
2. Incidence of which infection is increasing
3. How is HSV -1 transmitted
4. HSV infections may initiate what (2) things
1. Neutralizing antibodies
- half develop recurrent disease sometime during their life
2. Incidence of HSV2 increasing
- common venereal disease
- transmitted by INTERCOURSE (lots peeps having sex)
- associated with CERVICAL CARCINOMA; DNA tumor virus, may potentiate HPV in causing cervical cancer
3. HSV-1 is transmitted by SALIVA
- contaminated eating utensils
- low SES - low hygiene condition will increase the spread
4. HSV may initiate;
A. Erythema multiforme; most cases preceded by recurrence of HSV infection
B. Idiopathic neuropathic; Bell’s palsy, trigeminal neuralgia, temporal lobe epilepsy
36
1. What xracteristic of HSV infected cells can be detected by a TZANCK SMEAR ***
2. What will you see in cell culture?
- which is best for isolation of virus
- what temperature for transport?
3. Serology?
4. PCR?
1. Tranck smear
- scrape cells from base of lesion
- stain and look for NUCLEAR INCLUSIONS
- EM (electron microscope) is not specific
2. VIRUS ISOLATION in cell culture
A. Vesicular fluid is best for isolation of virus
B. Virus is very heat sensitive so take care in transport to lab (4degree)
3. Serology
- neutralization (antibody reduces infectivity of virus as determined by reduction in number of plaques)
- FLUORESCENT antibody attached to viral proteins on or in infected cells show cells are infected
4. PCR
- PCR based methods available for HSV encephalitis
37
Identify major treatment options for HSV and limitations (5)
1. Chemotherapy
2. NUCELOSIDE ANALOGS
A. Acyclovir (ACV); acycloguanosine
- oral ACV reduces severity of recurring disease
- inhibits viral DNA polymerase (early protein) and acts as a chain terminator
- ACV is inactive when administered
- ACV must be prophorylated by VIRAL THYMIDINE KINASE and then cellular kinases to form triphosphate
B. Valacyclovir
- taken orally and has better absorption xracters
- converts to ACV
C. Penciclovir
- must be phosphorylated for activation
- has longer life than ACV
3. Ganciclovir; more toxic than ACV
4. TFT; trifluorothymidine
5. Foscarnet and PFA (phosphonoformic acid)
- inhibits viral DNA polymerase
- foscarnet resembles diphosphate group and bind to polymerase
**Drug resistant mutation can occur
38
Relationship between varicella and zoster
1. Varicella (chicken pox) ; primary infection
- infection occurs in seasonal epidemics as chicken pox (varicella)
- contracted from another infected individual, usually a child
- systemic infection resulting in a generalized vesicular rash
2. Herpes Zoster (shingles); latent infection
- asymptomatic with no virus or virion proteins produced
- viral DNA resides in the cells of DORSAL ROOT GANGLIA (DRG)
3. Varicella zoster (shingles) ; recurrent infection
- virus travels down the sensory nerve fiber and infects epithelial cells inundated by the fiber
- infections are unilateral, painful vesicular eruptions localized to the dermatome, usually in the head or upper trunk
- severe systemic infections observed in immune suppressed individuals
39
Identify disease based on presentation
- Successive crops of lesions in a specific area of skin (asynchronous crops of lesions)
**how different from another condition with synchronized lesion in specific site?
1. Varicella (chicken pox) primary infection
- small pox however has;; one crop of lesion sin a specific area of skin (synchronized)
- Complications; Adults - pneumonitis (X ray show calcified nodules), Children - encephalitis (male has 2 times greater incidence than female), hemorrhagic varicella, bleeding in vesicles, GUT, GU
- Immunologically compromised and non-immune neonates; it is severe with 20% mortality. AIDS Patient
- REYE SYNDROME; aspirin related - hepatic failure, encephalopathy
40
Identify disease based in presentation
a. Occurs only with history of varicella infection,
verified by specific antibodies in serum
b. Second attacks unusual, but may occur
c. 1% of population annually
d. Resides in (dorsal root) ganglia of
affected nerve--recurs at dermatomes, thoracic
(50%), ophthalmic (15%), lumbar (15%), cervical
(10%)
e. Subject usually over 50 years
Herpes zoster (recurrent infection)
41
Evaluate risk of virus transmission to other patients from a patient with varicella vs zoster
1. Varicella (primary infection)
- RESPIRATORY ROUTE (mucosa) - VIREMIA - SKIN - macular popular, vesicular, non umbilicated - crusts not infectious, (each crop of lesions arise in asynchronous progression)
- 2 week incubation period - fever with each crop - widespread rah produce high fever (spread from trunk to face)
* *fever not as high as rash
2. Zoster
- occur only with history of varicella infection
- zoster patient can be source of chicken pox epidemic due to virus in vesicular fluid
- **Zoster can be activated via; X-rays, CNS tumor, trauma, immunosuppression (AIDS), exposure to CP or zoster - all may reactivate VZ virus
42
1. Pooled gamma globulin vs zoster-immune globulin in preventing varicella in exposed individuals
1. Pooled gamma globulin
- from patients convalescing from zoster
2. Zoster-immune globulin
- prevention of complication
* *given IM to immunocompromised patients within 2-3 days of exposure to VZ
* Antibody neutralizes Varicella zoster virus
43
What vaccines used for varicella/zoster
**what is approved for treatment of immunosuppressed patients
*Attenuated live vaccine which is recommended for routine vaccination of children - 2 doses recommended. Dose 1 at 12-15 months, dose 2 at 4-6 yrs and for susceptible older children, adolescents and adults
1. 2 varicella vaccines available
A. Single antigen vaccine (VARIVAX)
B. Combination vaccine - MMR (MMRV or pro quad)
2. Zoster vaccine
- Zostavax; individual over 60 years; one immunization. Reduce likelihood of shingles by 50%
- Shingrix-recombinant; given in 2 immunizations
* *Adenine arabinoside- approved for treatment of immunosuppressed patients with V or Z
* *Oral ACV also recommended
* *IV ACV for immunocompromised children
44
Describe 3 clinical entities associated with CMV
**how does CMV defy a rule of virus
CMV classic
1. “Owl eye” inclusion
- seen by microscopy of HE stain
2. Perinuclear cytoplasmic inclusions
3. Cell Enlargement
* *CMV is an exception to the rule of virus nucliec acid in that it has both DNA and RNA
- Ds DNA
- viral mRNA
45
Implications of CMV infection in;
1. Patients who have undergone open heart surgery
2. Been given multiple blood transfusions
1. Open heart surgery
- CMV infection is major cause of transplant failure (kidney)
- new organ might have CMV
- more severe infection if patient is immunosuppressed by drugs
3. Blood transfusion MONONUCLEOSIS
- 3-4 weeks after transfusion (lymphocyte in transfusion carry CMV)
**Remember that CMV and EBC initiate infections by infecting cells of respiratory system and regional lymph nodes - then spread to secondary sites of infection
46
1. Significance of high incidence of CMV in patieths with; Immunologic deficiency, Disseminated malignancy, Those receiving steroid therapy
2. Disease possibility (5)
1. In humans LATENT INFECTIONS can be established in MONOCYTES AND CERTAIN BONE MARROW CELLS - reactivated by immunosuppression (HIV)
2. Disease posssibilities;
A. congenital infection (TORCH); primary infection in mom (but asymptomatic, virus in urine) - CMV infects fetus in utero
B. perinatal infection; acquired newborn infection
I) birth canal; last trimester CMV secreted in cervix - due to reactivation of latent infection.
II) Newborn acquire from mother’s milk; mostly asymptomatic unless newborn is premature or immunocompromised
C. infection of normal (not compromised) children and adults
- mostly asymptomatic
- saliva, urine, treats, blood serve as vehicles for viral transmission
- sex in adults; semen and cervical secretions have virus or virus infected lymphocytes
D. receipients of transplantation and transfusions
E. infection of immunocompromised individuals (HIV cause retinitis)
47
What is role of EBV is initiating various cancers
EBV and cancer
- Epstein Barr virus is associated with lymphomas (e.g endemic burkitt lymphoma)
48
Distinguish btw EBV and CMV mononucleosis
Mononucleosis
1. EBV; heterophile antibody POSITIVE
- EB virus is main cause of mononucleosis
2. CMV; heterophile antibody NEGATIVE (on monospot)
49
What is torch?
**Of the torch infections which virus is most common cause of congenital disease
TORCH - major cause of congenital infections
A. Toxoplasmosis - Toxoplasma gondii - protozoan
B. Other--Treponema pallidum syphilis, listeria--gram + rod
(Listeria monocytogenes)
C. Rubella
D. Cytomegalic inclusion disease
E. Herpes simplex virus
**CMV most common virus that cause congenital infection. Rubella not as common due to vaccine
50
Picornaviridae
1. Major structural features of the virion
- strandedness?
- RNA type
- 5’ end? 3’ end?
- capsid? Component of capsid? Envelope?
2. Prototype and other example
1. Structure
A. Single stranded, non segmented RNA
B. (+) RNA; + polarity genome can act as messenger RNA. Genome (AUGGC) can “transfect” cells - naked genomic RNA is introduced into a cell and infection is initiated
C. RNA has poly A at 3’ end and VPg at 5’ end (VPg is not a cap struture).
**VPg is a viral protein g
D. Capsid has 4 structural proteins; VP1, VP2, VP3, VP4. Icosahedral capsid (no envelope)
E. Capsid structure is acid stable (enterovirus) and also stable to the environment
F. Capsid structure is NOT acid stable (rhinovirus)
2. Classes
A. Enterovirus; polio virus, coxsackievirus grp A and B, echovirus, hepatitis A
B. Rhinovirus; RSV?
51
Picornaviridae
Stages of infectious cycle
1. Attachment
- to what receptor?
- what capsid protein do you lose?
2. Uncoating
- occurs where?
- what capsid proteins removed?
3. (+) RNA
- what is lost for translation to occur
- what Association must occur for translation to occur
- what end of virus mRNA facilitates translation
4. Translation
- what is formed
Stages of infectious cycle
1. ATTACHMENT; to cell Nectin receptor - for PVR or CD155 immunoglobulin superfamily
- lose VP4 protein from capsid - penetration by endocytosis (viropexis)
2. UNCOATING; occurs in cytoplasm (site of viral replication)
- VP1, VP2, VP3 are removed from the virus particle to release the RNA
3. Viral genome is (+) polarity RNA; can serve as a messenger RNA
- following loss of VPg, the genomic RNA can be translated into viral proteins
- **genomic RNA associates with cellular ribosomal subunits for translation to occur
- The 5’ end of viral mRNA consists of specialized structure that facilitates translation
- IRES (internal ribosome enters site) consists of about 700 ribonucleotides and is essential for viral mRNA translation
4. TRANSLATION
- viral polyproteins (P1, P2, P3); linked together as one continuous large protein
* *(+) RNA w/o VPg is translated into polyprotein P123 - precursor protein
- P123 is proteolytically cleaved into both structural polypeptides and enzymes via proteolytic cleavage steps
52
Picornaviridae
Mechanism of viral RNA replication (describe diagrams)
Mechanism of replication
1. Polyprotein P123 - P1, P2, P3 (proteolytic proteins)
```
2.
A. P123 - P1
- P1 - VPo - VP2, VP4
- P1 - VP1
- P1 - VP3
** all capsids (no replicated genomes yet)
```
B. P123 - P2
- P2 - proteases
C. P123 - P3
- P3 - VPg (needed for replication)
- P3 - 3D (RNA dependent RNA polymerase)
• 3D associates with host factor that copy (+) strand into (-) strands (called RI - replication intermediate). Copies the template and then copies the copy (replication intermediate)
• End result; ASSEMBLY - poliovirus is lytic so cause cell to burst open
53
Picornavirirdae
Mechanism of viral RNA replication
A. What is required for replication
B. What 2 is then involved? What forms RI?
C. VPO is proteolytically cleaved to what (2) during maturation
Replication; occurs in the cytoplasm
A.
(+) RNA with VPg = RNA for replication
(+) RNA without VPg = mRNA that is translated
B. Replication; viral RNA polymerase (3D) + Host factor (Hf)
- RNA polymerase and genomic RNA with VPg at its 5’ end is then involved in viral RNA replication
- A replicative intermediate (RI) is formed due to the + genomic RNA being converted to many - RNAs; then full length - RNA is copied into + genomic RNAs (another RI)
C. VPO proteolytically cleaved to VP2 and VP4 during viral maturation
54
Spectrum of disease caused by members of picornaviridae
- Assembly of infectious virus particles + production of empty capsids protein produced in excess); note empty capsids are not infectious; cell lysis occurs as an end result of infection
- At the end of a replication cycle each cell can yield approximately 10,000 virus particles
**Picornavirus; HEPATITIS A, colds, meningitis, poliomyelitis
55
Antivirals
Summary of agents
1. Anti- herpetic
- 3 major first line
- 1 tropical
2. Anti-CMV
- 2 first line
- 2 second line
3. Anti-influenza
- 4 total
- which is the best and why?
4. Hepatitis B/C
- 3 major
- what increase duration of action?
1. Anti-herpetic (HSV, VZ)
A. First live (AVF), Acyclovir, Vancyclovvir, Famciclovir
B. Topical; penciclovir
2. Anti-CMV
A. First line (fewer A.E); Ganciclovir and Valganciclovir
B. Second line; Cidofovir and Foscarenet
3. Anti-influenza
- Ostelmavir (tamiflu), amantadine, rimantadine, zanamivir
- Ostelmavir is most preferred because it covers both types of influenza
4. Hepatitis B/C
- Interferons +/- ribavirin, lamivudine
- pegalated to increase duration of action
56
Identify the following antivirals based on the MoA
1. Inhibition of viral genome replication (2)
2. Inhibit viral un-coating
- block M2 (pH gated proton channel) that opens in response to acidification (2)
3. Inhibition of viral release
- Neuroaminidase cleaves Sialic acid from the membrane glycoproetins releasing the virus (2)
1. Inhibit viral replication
- Nucleoside and Nucleotide analogues (Acyclovir, Ganciclovir)
2. Inhibit viral in-coating
- Amntadine/Rimantidine
3. Inhibit viral release
- Ostelmivir and Zanamivir
57
Identify 5 families of DNA viruses
DNA virus
1. Poxvirus (smallpox)
* 2. Herpesvirus (CHICKEN POX, SHINGLES, ORAL and GENITAL HERPES)
3. Adenoviruses (sore throat, conjunctivitis)
* 4. Hepadnavirus (HEPATITS B)
5. Papillomavirus (Warts)
58
Identify 9 families of RNA viruses
1. Rubella (German measles)
2. Rhamdoviruses (rabies)
* 3. Picornavirus (Hepatitis A, colds, meningitis, poliomyelitis)
4. Arenaviruses (Lassa fever, meningitis)
* 5. Flaviviruses (Hepatitis C, West Nile meningoenchpalitis, yellow fever)
6. Orthoxymyxovirus (influenza)
7. Paramyxoviridae (Measles, Mumps)
* 8. Cornovirus (SARS, colds)
9. Retrovirus (HIV)
59
Identify common viral infections (6)
1. Herpes Family; varicella (chickenpox), Zoster (shingles), encephalitis
2. CMV; common in transplant patients (retinitis and pneumonia)
3. Influenza; types A and B (ages 15-30 most common)
4. Hepatitis B and C
5. RSV - respiratory syncytial virus
- pneumonia in children
6. HIV
- systemic destruction of helper T cells
60
Identify drugs used in treatment of herpes (HSV 1 and 2), zoster. (1 of 4)
- MoA
- prodrug vs drug
- Use
- Bioavailability
- Adverse effects
1. ACYCLOVIR (drug)
A. MoA; acyclovir is phosphorylated to monophospahte (thymidine kinase of virus) - diphosphate (cellular kinase) - triphosphate (cellular kinase)
B. Use
- herpes simplex (cold sores); 1 episode require 10 day treatments, recurrent episode require 5 day treatment
- encephalitis; IV acyclovir is DOC. Good CSF penetration for treatment of viral meningitis
C. Bioavailability; poor (10-30%)
- available as IV, PO (5x daily damn!!) and topical
D. Adverse effect; renally eliminated so require dose reduction
- reversible NEPROTOXICITY; crystalline nephropathy more likely with high dose bolus vs IV formulation. **Please slow IV infusion
- CNS (HA, hallucinations and seizures)
61
Identify drugs used in treatment of herpes (HSV 1 and 2), zoster. (2 of 4)
- MoA
- prodrug vs drug
- Use/uniqueness
- Bioavailability
- Adverse effects
2. Valacyclovir (Valtrex); Prodrug
A. MoA
- valacyclovir - acyclovir (intestinal and hepatic metabolism) - monophosphate (viral thymidine kinase) - di and then triphosphate (cellular kinase)
B. Use ; P.O. only
- HSV, zoster,
- better bioavailability so don’t require as frequent dosing as acyclovir
- **best outcome if used within 48-72 hours of lesion/rash
C. Bioavailability ; 70%
- P.O. route reach similar levels as IV doses of acyclovir
D. Adverse effects
- CNS; HA, tremors, hallucinations
- N/V/D; dose and age dependent
- Hepatic; hyperbilirubinemia
62
Identify drugs used in treatment of herpes (HSV 1 and 2), zoster. (3 and 4)
- MoA
- prodrug vs drug
- Use/uniqueness
- Bioavailability
- Adverse effects
3. Penciclovir (Denavir)
A. MoA; similar to acyclovir
- penciclovir - monophosphate (thymidine kinase) - di and then triphosphate (cellular kinase)
B. Use; TOPICAL CREAM
- herpes simplex cold sore treatment
C. Adverse effect ; application site irritation
4. Famciclovir (famvir); PRODRUG FOR #3
A. MoA; similar to above
- Famciclovir - penciclovir (intestine and hepatic enzyme) - monophosphate (thymidine kinase) - di and tri phosphate (cellular kinase)
B. Use
- Herpes simplex/zoster
- best results if started w/in 72 hours
C. Bioavailability; 65-77%
D. Adverse effect; renally excreted
- Headache and GI
63
1. What are the top 3 antiherpetic?
2. The 4th one is back seat becuase it is only available as topical cream - which is this?
- identify other topical antivirals
1. 3 antiherpetic (AVF)
- Acyclovir (P.O., IV, topical)
- Vanciclovir (P.O. only)
- Famciclovir
- Penciclovir (topical)
2. Other topical anti-virals
- Docosanol (Abreva); oral herpes simplex
- Imiquimod (Aldara); genital/perianal warts
- Podofilox (Condylox); genital/perianal warts
- Podophyllin; 25% liquid administered in clinic directly to wart
64
Identify top 2 treatment for CMV
- MoA
- bioavailability
- Adverse effects
- treatment
1. Ganciclovir (Cytovene)
A. MoA; same as acyclovir
- 10x more potent than acyclovir against CMV (in-vitro studies)
B. Bioavailability; 6-9%
- IV, Intravitreal (eye injection) and P.O.
C. Adverse effects
- **CNS with behavior changes and headaches
- N/V
- neutropenia (15-40%) and thrombocytopenia (5-20%)
D. Treatment
- CMV RETINITIS prophylaxis and treatment *****
```
2. Valganciclovir (Valcyte); PRODRUG
A. MoA; same as above
B; bioavailability
- better
C. Adverse effect; same as above
D. Treatment
- CMV treatment; use BID
- CMV prophylaxis; DAILY
**Intravitreal Treatment; surgically implanted to treat CMV retinitis in AIDS patients
```
65
Identify 2 second line antiviral for treatment of CMV
- MoA
- indication for use
- adverse effect
**which do you used if immunocompromised pt with HSV is resistant to ASV
1. Cidofovir (Vistide)
A. MoA; different from ACV
- interacts with DNA polymerase either as; alternative substrate or competitive inhibitor
B. Indication for use
- reserved for CMV retinitis for treatment failure of ganciclovir and/or foscarnet
- Administer with Normal saline to decrease chances of nephrotoxicity
- for CMV treatment, must be administered with PROBENECID
C. Adverse effect
- Highy nephrotoxic
- neutropenia
2. FOSCARNET (foscavir)
A. MoA; competes for pyrophosphate in viral DNA polymerase
- foscarnet is an inorganic phosphate that doesnt require phosphorylation by thymidine kinase
B. Clinical use (2nd line agent)
- CMV in AIDS patient (combi with GCV in recurrence)
* drug-drug interaction with zidovudine
- ACV resistant HSV in immunocompromised pts ***
66
Identify gold standard treatment therapy for hepatitis B/C
- MoA
- adverse effects
- clinical use
** gold standard treatment
1. Interferon
A. MoA; not understood
- available naturally (body produce it) or external source (injection/infusion)
*Pegalated (polyethylene glycol) to improve pharmacokinetic parameters after injection
B. Adverse effect
- flu like symptoms; fever, nausea, ache, sore muscles, joint pain and fatigue
- thrombocytopenia, granulocytopenia
- neurotoxicity (depression, confusion)
- alopecia
- personality changes (common in kids)
- Monitor LFTs
C. Clinical use
I) Type I interferons
- Alpha; Hep B, hep C, condylomata acuminata (HPV), kaposi’s sarcoma by herpes virus
- Beta; treat MS
II) Type II interferons
- gamma (typically produced by T cells of immune response)
- not commercially made
67
Identify therapy for hepatitis B
**has black box warning ; lactic acidosis and severe hepatomegaly
- MoA
- bioavailability
- adverse effect
- treatment
2. Lamivudine (Epivir)
A. MoA; inhibits reverse transcriptase
B. Bioavailability - 80-90%
- renally eliminated
C. Adverse effect
- BLACK BOX WARNING; lactic acidosis and severe hepatomegaly (fatty liver steatosis)
- HA, fatigue, insomnia, peripheral neuropathy**
- arthralgias and rash
D. Treatment options
- combo with interferons to treat hepatitis B
- PART OF HIV TREATMENT REGIMEN
68
Identify therapy for Hep C
- very long half life (200-300 hours at steady state)
- treat RSV
- MoA
- adverse effect
- treatment
Ribavirin (Rebetrol).
A. MoA; unknown
*touted as broad spectrum antiviral
B. Adverse effect; renally eliminated
- dose related anemia (may require dose reduction or epgoen
- combo with interferons increases anemia, fatigue, depression
C. Uses
- tract hep C with interferon
- RSV bronchiolitis and pneumonia in hospitalized children
69
Identify topical treatment for warts (condylomata acuminate)
MoA
What to keep in mind
Imiquimod (Aldara)
MoA; induces local immunogen-response (interferon alpha, beta, gamma and TNF) to DECREASE VIRAL LOAD
**Keep in mind; recurrence is common
70
1. Identify therapy for influenza A and B
- best treatment for flu
A. MoA
B. Use
C. Adverse effects
2. Identify therapy for flu A and B
- cause WHEEZING AND BRONCHOSPASM
1. Osteltamivir (Tamiflu)
A. MoA; inhibit neuroaminidase which is required for release of virus from infected cells
B. Bioavailability; P.O. - cause GI side effect (N/V)
- take with food
C. Use
- influenza types A and B
```
2. Zanamivir
MoA; same as osteltamiir
**Adverse effect; How to use; dry powder delivered by inhalation which may cause Wheezing and BRONCHOSPASM
- avoid in COPD and Asthma patients
Use; influenza type A and B
```
71
Identify 2 second line therapy for influenza virus
- which is last ditch effort for late Parkinson’s
- what type of influenza do both treat
- why isn’t it first line?
- which is safe in elderly ‘
- which require renal adjustment
1. Amantadine
- Amantadine is last ditch effort for LATE PARKINSONS
- renal adjustment required for amantadine
- Adverse effects more pronounced; CNS (confusion, insomnia)
2. Rimantadine
- no renal adjustment
- safe in elderly patients
* Both TREAT AND PREVENT INFLUENZA A; A is more prevalent than B
* CDC now advise not to use this as first line drug for flu due to resistance
** MoA; prevents virus from entering host cell
• baseball hat with things coming off the sides
72
# Define inflammation
*Immunology vs pathology
Inflammation; vascularized tissue response to infection and damaged tissues
A. Immunology; protective process
B. Pathology; can damage tissue
73
Describe general features of inflammations (5)
Features of inflammation
1. Offending agent is located in extravascular tissues and is recognized by host cells and molecules
2. Both vascular reaction and cellular response are derived from plasma proteins and various cells. Leukocytes and plasma proteins are recruited from the circulation to site where offending agent is located
3. Leukocytes and proteins work together to destroy/eliminate offending substance
4. The reaction is controlled and terminated
5. The damaged tissue is repaired
74
Describe 2 major components of inflammation
**Identify cardinal signs of inflammation (5)
2 components
1. Vascular reaction
A. Vasodilation; initial vasoconstriction (pallor) and then vasodilation.
B. Vascular leakage and edema; increased vascular permeability. Loss of intravascular fluid lead to blood flow stasis and exudate (accumulation of fluid) in interstitial tissues and/or body cavities. Account for swelling, pain and loss of function
**NOS promote vasodilation (iNOS)
2. Cellular response; leukocyte emigration and extravascular tissues
A. Migration and rolling (selectins - E,L,P)
B. Activation and adhesion (adhesions)
C. Transmigration (chemotaxis - CD31 or PECAM-1)
**NOS inhibit cellular response by inhibiting recruitment
**Pain, redness, heat, swelling and loss of function - cardinal signs of inflammation
75
Define and compare transudate and exudate
1. Transudate; ultrafiltrate of plasma with no increase in vascular permeability
- low protein content (mostly albumin)
- low specific gravity
- little or no cell material
- from osmotic or hydrostatic imbalance
2. Exudate
- high protein content
- cellular debris
- from tissue injury or ongoing inflammation
76
Describe the leukocyte recruitment, extravastation, and phagocytosis in acute inflammation
1. Leukocyte recruitment
A. Margination, rolling (selectin) and adhesion (INTEGRINS) to endothelium
B. Migration across the endothelium and vessel wall
C. Migration in the tissues toward a chemotactic stimulus (chemotaxis)
2. Extravastation (diapedesis by chemotaxis)
3. Phagocytosis; of microbe into phagosome - kill microbes. Phagocytosis and intracellular killing are important for destruction of microbes. 3 step process ;
- recognition and opsonization (by IgG opposin)
- engulfement (form vacuole)
- killing/degradation ; by ROS and NO or by lysosomal enzymes in phagolysosome. The H2O2-MPO-halide system is the most efficient bactericidal system of neutrophils
77
1. Identify chemical mediators of inflammation - exogenous
| 2. List and discuss the role of cell and plasma derived mediators of inflammation (Endogenous mediators)
1. Exogenous
A. Bacterial endotoxin (LPS)
B. Bacterial peptides (N-formyl methionine)
2. Endogenous
A. Cell derived mediators ; sequestered in intracellular granules
I) preformed mediators in secretory granules; Histamine (from mast cell), serotonin (from platelets), lysosomal enzymes (neutrophils, macrophages)
II) newly synthesized; prostaglandins, leukotrienes, platelet activating factors, activated oxygen species, NO, cytokines
B. Plasma derived mediators (liver); typically circulate in inactive form and undergo proteolytic cleavage to acquire their biological activities
I) Factor XII (Hageman factor); kinin system (bradykinin - pain), coagulation/fibrinolysis system
II) complement activation; C3a and C5a (anaphylatoxins), C3b, C5b-9 (MAC)
78
Compare and construct acute vs chronic inflammation
A. Onset
B. Cellular infiltrate
C. Tissue injury, fibrosis
D. Local and systemic signs
1. Acute inflammation; innate immunity
A. Onset; fast - minutes or hours or few days
B. Cellular infiltrate; PMN (neutrophils)
C. Tissue injury, fibrosis; mild and self-limited
D. Local and systemic signs; prominent (increased vascular permeability - vascular leakage
2. Chronic inflammation; adaptive immunity
A. Onset; slow (days)
B. Cellular infiltrate; monocytes/macrophages and lymphocytes
C. Tissue injury, fibrosis; severe and progressive
D. Local and systemic signs ; less prominent - may be subtle
79
Describes morphologic patterns of acute inflammation
1. Low MW protein (albumin), no cells - blister
• E.g burn or viral infection
2. Larger protein - no cells
3. Local defect of organ surface produced by necrosis and inflammation
4. In addition to albumin, fibrin and other proteins, the exudate contains neutrophils
5. Occur in Type I hypersensitivity (allergic) reactions and certain parasitic infections
Acute inflammation ; based on type of exudate
1. Serous inflammation
- serous effusion (clear yellow fluid , no cells, small MW protein mostly ALBUMIN)
- from skin blister of second degree burn or viral infection (varicella, HSV), repetitive trauma
2. Fibrinous inflammation ; larger protein - FIBRIN
- fibrinous is different from fibrous
- E.g fribinous percarditis in rheumatic cardinitis
- Fibrinous pneumonia (pneumonitis) in chlorine gas inhalation or viral infection e.g influenza
3. Ulceration; occur when tissue necrosis and resultant inflammation exist on or near a surface.
- occur in mucosa of mouth, stomach, intestines, GI tract
- in subcutaneous tissues of the lower extremities in older persons e.g peptic ulcer of the stomach or duodenum in which acute and chronic inflammation coexist
4. Purulent (suppurative); exudate composed of fluid, protein and dead/dying neutrophils and other cells is pus
- suppurative pneumonia or meningitis due to pyogenic infection (neisseria meningitis)
- Pyosalpinx – purulent infection of the fallopian tube e.g. due to Neisseria gonorrhoeae.
5. Eosinophilic exudate; eosinophil is predominant in exudate
- asthma, allergic rhinitis, nematode infection, ascariasis of small intestine
80
Describes morphologic patterns of acute inflammation
6. Damage to endothelial cells and vessel walls allow RBCs to leak into surrounding tissue
7. Diffuse area of acute inflammation composed of edema fluid, bacteria, and neutrophils spread through tissues e.g strep pyrogens
8. Injury results in necrosis of affected tissue lining a surface - form membrane that composed of fluid, proteins and neutrophils, RBC and necrotic tissue
9. Inflammation in tissue that contain mucin-secreting gland and likely stimulate mucus
10. Acute inflammation composed of pyogenic exudate and necrotic tissue
6. Hemorrhagic; RBCs leak into tissue
- Rickettsia prone to damaged endothelial cells
- e.g Rocky Mountain spotted fever
7. Cellulitis ; edema fluid, bacteria and neutrophils
- skin and subcutaneous tissue e.g strep pyogenes. (Necrosis not normally present)
- flesh eating infections (necrosis present)
8. Pseudomembranous
- pseudomembrane obstructing the upper airway in diphtheria
- pseudomembranous enterocolitis in patient treated with clindamycin
- many exudate combined - fibrinopurulent or mucopurulent
9. Mucinous ; bronchitis
10, Abcess; although acute - can be present for a long time but eventually get walled off by fibrous connective tissue
- furuncle ; boil - subcutaneous abcess near hair follicle (S. Aureus)
- carbuncle; coalesced furuncles
81
1. Define granulomatous inflammation and list the key inflammatory cells involved
2. Types of granuloma
1. Granuloma inflammation - type of chronic inflammation
A. MACROPHAGES and lymphocytes
B. EPITHELIOID CELLS***; activated macrophages (by gamma interferon)**
C. Presence or absence of giant cell ; fusion of more than one epithelioid cells
2. 2 types granuloma
A. Foreign body type; giant cell
B. Immune granulomas; langhans giant cells
- Non-caseous (Sarcoidosis); react to foreign material (Talc or suture) - crowns, cat scratch disease
- caseous (TB and fungus); tissue necrotic pattern
82
1. Describe systemic effects of inflammation
| 2. Non specific indicators of inflammation (lab tests) - 3
1. Systemic Effects of inflammation
A. Bacterial products; LPS (exogenous pyrogens)
B. CRP (C-reactive protein); increased risk of MI or stroke
C.
- Neutrophilial (bacterial infection)
- Lymphocytosis (viral infection)
- eosinophilia (bronchial asthma, hay FEVER and parasite infestation
2. Nonspecific indicators of inflammation
A. Left shift ; **LEUKEMOID REACTION. More immature neutrophils (bands) than mature neutrophils in peripheral blood smear indicate bacterial infection
B. ESR; the increase in amount of fibrinogen and erythrocyte sedimentation rate indicate an inflammatory response
C. C-reactive protein ; CRP is a glycoprotein synthesized by the liver. Blood levels of CRP elevated in an inflammatory response
83
Describe deficiencies in inflammatory response
1. Autosomal Disease. Seen in infants and children. Neutropenia with recurrent infections. Oculcutaneous albinism. Aberrant granules inn neutrophils and other WBCs. Melanocytes. Basic defect is unknown - may be in microtubules
2. X linked - mainly in boys. Basic defect in NADPH oxidase 0 deficiency of ocygen dependent generation of H2O2 and microbial killing
* * susceptibility in what organisms
Deficiencies in inflammation
1. Chediak-Higashi disease
2. CGD (chronic granulomatous disease)
- recurrent infections especially by catalase - producing microbes
**
C- candida
A- staph Aureus
T
A- Pseudomonas Aeruginosa
L- listeria
A- Aspergillus
S- Serratia
E- E. Coli
84
Chemical mediators of inflammation
1. Identify all cell-derived mediators (9)
2. Identify plasma protein-derived mediators (2)
**also list their sources and functions
1. Cell-derived
A. Histamine; from mast cells, basophils, platelets
- VASODILATION, increased vascular permeability, endothelial activation
B. Serotonin; platelets
- VASOCONSTRICTION
C. Prostaglandin; mast cells, leukocytes
- vasodilation, PAIN*, fever
D. Leukotrienes; mast cells, leukocytes
- increased vascular permeability, chemotaxis, leukocyte adhesion and activation
E. PAF (platelet activating factor); leukocyte, mast cells
- vasodilation, increased vascular permeability, leukocyte adhesion, chemotaxis, degranulation, oxidative burst
F. ROS (reactive oxygen species); leukocytes
- killing of microbes, tissue damage
G. NO (nitric oxide); endothelium, macrophages
- vascular smooth muscle relaxation, killing of microbes
H. Cytokines (TNF, IL1, IL6); macrophages, endothelial cells, mast cells
- *Local; endothelial activation (expression of adhesion molecules)
- *Systemis; fever, metabolic abnormalities, hypotension (shock)
I. Chemokines; leukocytes, activated macrophages
- chemotaxis, leukocyte activation
2. Plasma protein-derived
A. Complement ; plasma (produced in liver)
- leukocyte chemotaxis and activation, direct target killing (MAC), vasodilation (mast cell stimulation)
B. Kinin; plasma (produced in liver)
- increased vascular permeability, smooth muscle contraction, vasodilation and pain
85
Chemical mediators
1. Identify 2 types of vasoactive amines
2. 2 types of lipid mediators
1. Vasoactive amines (stored as preformed molecules in cells and have important action in blood vessel)
A. Histamine; vasodilation, increased vascular permeability, endothelial activation
B. Serotonin ; vasoconstriction
2. Lipid mediators (cell derived - produced from Arachidonic acid present in membrane phospholipids)
A. Prostaglandins; vasodilation, PAIN, fever
- generated by COX 1 and COX 2
- aspirin and NSAIDs (Indomethacin, ibuprofen, naproxen) inhibit COX 1 and COX2 - reduce pain and fever
- steroids inhibits Phospholipase (inhibit Arachidonic acid so no prostaglandin or leukotrienes)
B. Leukotrienes ; increased vascular permeability, chemotaxis, leukocyte adhesion and activation
- steroids inhibit arachidonic acid
- not affected by steroids
86
Identify chemical mediator
• phospholipid mediator (acetyl glycerol ether phosphocholine).
• produced by neutrophils, monocytes, basophils, endothelium, platelets,
• Actions – platelet stimulation, vasoconstriction, bronchoconstriction,
others. vasodilation, increased vascular permeability, leukocyte activation (adhesion, chemotaxis, degranulation, oxidative burst).
• generated from membrane phospholipids by action of phospholipase A2.
Platelet-Activating Factor (PAF)
87
Chemical mediators - plasma derived
Identify the types
1. Increase vascular permeability and cause vasodilation by binding to mast cells and inducing histamine release (ANAPHYLATOXINS) thereby increase vascular permeability and cause vasodilation. They are called anaphylatoxins because they have effects similar to those of mast cell mediators that are involved in the reaction called anaphylaxis
2. CHEMOTACTIC for neutrophils, monocytes, eosinophils, and basophils. Increases adhesiveness of neutrophils to endothelium. Stimulates synthesis and secretion of arachidonic acid metabolites.
3. Opsonization & Phagocytosis: binds to microbial surface and
promotes phagocytosis.
**Identify biological effects of complement fragments
1. C3a, C5a ; anaphylatoxins
- chemotactic, stimulate immune response
- suppress immune system
- mast cell stimulation
- activation of tissues (Cyclooxygenase and lipoxygenase)
2. C5a
3. C3b and iC3b
** Biologic effects of complement fragments
- Vascular phenomena: (C3a, C5a; anaphylotoxin)
• Vasodilation
• Increase vascular permeability
- Leukocyte adhesion, chemotaxis and activation: C5a (chemotaxin)
- Phagocytosis: C3b and iC3b Opsoninaztion
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1. Identify
- It is a protein synthesized by the liver
- called clotting factor XII( inactive form)
- Provides additional source of vasoactive mediators
- Activated by: Negatively charged surfaces , Bacterial LPS, sodium urate crystals and enzymes (e.g. Trypsin and Plasmain)
- **activate both clotting system and kinin system
2.
- vasoactive peptides derived from plasma proteins kinnogen
- It is a vasodilator, increases vascular permeability, bronchial smooth muscle contraction, pain.
- It is short-lived
1. Hageman factor
| 2. Kinins
89
In chronic inflammation;
1. What activates macrophages?
2. What are the 2 pathways of macrophage activation?
- functions of each pathway
1. Gamma interferon activates macrophages
2. 2 pathways
A. Classically activated macrophage (M1)
I) ROS, NO, lysosomal enzymes; microbicidal actions - phagocytosis and killing of many bacteria and fungi
II) IL1, IL12, IL23, chemokines; inflammation
B. Alternatively activated macrophage (M2)
I) Growth factors TGFbeta; tissue REPAIR, fibrosis
II) IL10, TGFbeta; anti-inflammatory effects
**Activated macrophages in chronic inflammation has same mechanism as acute ; migration, rolling etc
90
How do macrophages and lymphocytes interact in granuloma formation
BI-DIRECTIONAL WAY ; these interactions play an important role in propagating chronic inflammation
Macro- phages display antigens to T cells, express membrane molecules (called costimulators), and produce cytokines (IL-12 and others) that stimulate T-cell responses.
Activated T lymphocytes, in turn, produce cytokines, described earlier, which recruit and activate macrophages (classic pathway), promoting more antigen presentation and cytokine secretion. The result is a cycle of cellular reactions that fuel and sustain chronic inflammation.
91
Identify the following cytokines used in acute vs chronic inflammation
- principal sources
- principal actions in inflammation
**which cytokine is present in both acute and chronic (Remember that all the cytokines listed as acute can also play role in chronic)
1. Acute inflammation (5) ; TNF, IL-1, IL-6, chemokines, IL-17
A. TNF; macrophages, mast cells, T lymphocyte
- stimulates expression of endothelial adhesion molecules and secretion of other cytokines; systemic effects
B. IL-1; macrophages, endothelial cells, some epithelial cells
- similar to TNF; greater role in fever
C. IL-6; macrophages and other cells
- Systemic effects (acute phase response)
D. Chemokines; macrophages, endothelial cells, T lymphocytes, mast cells, other cell types
- recruit leukocytes to site of inflammation
- migration of cell in normal tissues
E. IL-17; T lymphocytes
- recruitment of neutrophils and monocytes
2. Chronic inflammation (3) ; IL-12, IFN-y, IL-17
A. IL-12; DC, macrophages
- increased production of gamma interferon
B. IFN-y; T lymphocytes, NK cells
- activation of macrophages (increased ability to kill microbes and tumor cells)
C. IL-17; T lymphocytes
- recruitment of neutrophils and monocytes
92
Identify the following principal mediators based on their roles in inflammation
1. Vasodilation (2)
2. Increased vascular permeability (3 groups)
3. Chemotaxis, leukocyte recruitment and activation (4)
4. Fever (2)
5. Pain (2)
6. Tissue damage (2)
1. Vasodilation (2)
- histamine
- Prostaglandin
2. Increased vascular permeability (3 groups)
- histamine and serotonin
- C3a and C5a (liberate vasoactive amines from mast cells)
- leukotrienes C4, D4, E4
3. Chemotaxis, leukocyte recruitment and activation (4)
- TNF, IL-1
- chemokines
- C3a, C5a
- leukotriene B4
4. Fever (2)
- IL1, TNF
- prostaglandin
5. Pain (2)
- prostaglandin
- bradykinin
6. Tissue damage (2)
- lysosomal enzymes of leukocytes
- ROS
