4. stem cells and regenerative medicine in the pancreas

Question 1

the biliary tree is a system of vessels that directs secretions from which three organs into what system?

Answer 1

secretions from the liver, pancreas and gallbladder are directed to the duodenum (the small intestine)

Question 2

secretions exit the biliary tree through a hole into the duodenum, what is this hole called?

Answer 2

Ampulla or Vater

Question 3

what type of organ is the pancreas?

Answer 3

a secretory organ

Question 4

what are acinar cells and what do they do?

Answer 4

they are exocrine ducts which secrete digestive enzymes

Question 5

what and where do pancreatic islets secrete?

Answer 5

hormones into the blood

Question 6

what are the two main hormones produced by these islets?

Answer 6

insulin and glucagon

Question 7

what do alpha cells produce?

Answer 7

glucagon

Question 8

what do beta cells produce?

Answer 8

insulin

Question 9

what is the function of the islets of Langerhans?

Answer 9

to regulate blood glucose homeostasis

Question 10

which cell type makes up 60-70% of the islets?

Answer 10

beta cells

Question 11

what is the function of insulin?

Answer 11

insulin lower blood glucose levels

Question 12

name three ways in which insulin functions?

Answer 12

stimulates the body cells to absorb glucose, the liver stores glucose as glycogen and fat cells convert glucose into fatty acids

Question 13

what is the function of glucagon?

Answer 13

raises blood glucose levels

Question 14

how does glucagon function?

Answer 14

stimulates the break down of glycogen stores in liver and muscle cells

Question 15

what is the range of normal blood glucose levels that needs to be maintained?

Answer 15

70-110mg/dL

Question 16

what happens when normal blood glucose levels are disturbed?

Answer 16

the pancreas will react in order to restore normal blood glucose levels

Question 17

what is type 1 diabetes classed as?

Answer 17

a chronic autoimmune disease

Question 18

what occurs in T1D?

Answer 18

T cells attack insulin secreting beta cells, this results in beta cells becoming unresponsive

Question 19

when does T1D manifest?

Answer 19

it usually manifest during childhood

Question 20

in terms of stem cell therapy, why will T1D be harder to treat than T2D?

Answer 20

a way to prevent T cells from attacking any new beta cells will need to be developed

Question 21

how many of the total cases of diabetes are T2D?

Answer 21

85-90%

Question 22

what is T2D?

Answer 22

a chronic, progressive disease characterised by an abnormal insulin action and secretion

Question 23

what is the initial problem seen in the development of T2D, which if controlled can prevent the development of T2D?

Answer 23

insulin resistance

Question 24

what is insulin resistance?

Answer 24

when key target tissues have diminished response to insulin

Question 25

what percentage of the pancreas is normally comprised for islet cells?

Answer 25

1-2%

Question 26

during insulin resistance, what happens to the islet?

Answer 26

beta cells increase in both size and number

Question 27

why do islet cells respond in this way to insulin resistance?

Answer 27

cells of the body aren't responding to insulin efficiently, and so the body is overcome the lack of responsiveness by producing more insulin

Question 28

what is glucose tolerance?

Answer 28

when blood glucose can be maintained during insulin resistance due to the increase in insulin production

Question 29

what happens after islet compensation in insulin resistance?

Answer 29

- there is impaired islet compensation/glucose tolerance - islets are overwhelmed and so beta cells begin to apoptose - number of islets decrease - reduction in number of beta cells per islet

Question 30

what is the result of insulin resistance and impaired islet compensation?

Answer 30

cells are less responsive to insulin in addition to the pancreas producing less insulin and type 2 diabetes manifests

Question 31

give three risk factors for T2D

Answer 31

- genetics - obesity - age

Question 32

during insulin compensation what percentage of the islets do beta cells make up?

Answer 32

90%

Question 33

what does the world heath organisation predict in relation to T2D? and what does this mean for pancreatic research?

Answer 33

- diabetes will be the 7th leading cause of death by 2030 | - the is a potential large financial for T2D research

Question 34

the liver, pancreas and biliary tree development starts at what day after fertilisation?

Answer 34

day 35

Question 35

describe the pancreas at day 35 after fertilisation?

Answer 35

their is a ventral pancreatic bud and a dorsal pancreatic bud

Question 36

how does the biliary system initially develop?

Answer 36

it develops as a tubular structure from the elongation and moulding of the posterior portion of hepatic diverticulum

Question 37

organs linked by the biliary tree are also linked during... ? and what does this tell us?

Answer 37

- development | - all of these cells are derived from a single multipotent population of cells

Question 38

what happens 40 days after fertilisation?

Answer 38

dorsal and ventral pancreatic bud rotate clockwise and fuse

Question 39

what forms at day 45 post fertilisation?

Answer 39

the biliary tree

Question 40

where is it common to find stem cells niches in this system?

Answer 40

associated with the biliary ducts

Question 41

where are stem cells found in the liver?

Answer 41

canal of hering at the tip of the biliary tree

Question 42

if the liver is damaged and hepatocytes are unable to respond, what may occur?

Answer 42

there may be streaming of stem cells from other regions

Question 43

what is the marker for biliary tree stem cells and their progeny (hepatic and pancreatic lineage)? and what can be put under control of this promoter for lineage tracing?

Answer 43

Sox9 | CreER

Question 44

what is the marker for cells of pancreatic lineage?

Answer 44

PDX1

Question 45

in linage tracing, what is floxed upstream of a reporter gene? and give an example of a reporter gene?

Answer 45

- stop codon | - lacZ

Question 46

when LacZ is expressed how can it be detected?

Answer 46

- addition of XGAL | - beta-galactosidase will hydrolyse the glycosidic bond and give rise to galactose and a blue product

Question 47

what cells will be marked by this lineage tracing? and what does this tell us?

Answer 47

- cells of the intestine, pancreas, liver and biliary tree | - sox9 marks a progenitor population within the biliary tree

Question 48

where are the multipotent progenitors of biliary tree?

Answer 48

peribiliary glands

Question 49

when these multi-potent progenitors decide to become pancreatic, what is their first fate decision?

Answer 49

whether to be ventral or dorsal

Question 50

what cell type becomes endocrine?

Answer 50

ventral progenitors

Question 51

these progenitors may become endocrine cells or...?

Answer 51

acinar cells

Question 52

give three possible fates of the multipotent progenitors of the biliary tree?

Answer 52

hepatic cells extra-hepatic cells pancreatic cells

Question 53

what two transcription factors determine where pluripotent stem cells will be endoderm?

Answer 53

sox17 and foxA2

Question 54

what two transcription factors drive endoderm cells to become pancreatic endoderm?

Answer 54

sox9 and pdx1

Question 55

which three cells types can the pancreatic endoderm then become? and what does this depend on?

Answer 55

- ducts cells, acinar cells, endocrine cells | - this depends on their gene expression

Question 56

in order to give rise to beta cells what do we need to drive cells to become?

Answer 56

endocrine progenitors

Question 57

altering what type of signalling in the body can change the fate between hepatic and pancreatic lineages?

Answer 57

non-carconical Wnt signalling

Question 58

how was this type of signalling that can change cells fate between hepatic and pancreatic lineages identified?

Answer 58

- hepatic and pancreatic progenitors were isolated by flow cytometry - RNAseq was used to define molecular identify of cells

Question 59

what is trans-differentiation?

Answer 59

when one committed cells gives rise to a different type of committed cell

Question 60

what is de-differentiation?

Answer 60

when a committed cells goes back to a less differentiated state

Question 61

how has acinar cells plasticity been shown in model animals?

Answer 61

following pancreatic duct ligation - where a ligature is made in the pancreas and a pancreatic duct is tied off - acinar cells can convert to embryonic type multipotent cells and give rise to acinar cells, duct cells and endocrine cells

Question 62

what does this animal model acinar cellular plasticity suggest?

Answer 62

that it is possible to take acinar cells and manipulate them to become beta cells

Question 63

under what conditions have interconversion between cells of the islet been observed?

Answer 63

experimental and pathological

Question 64

what is special about this interconversion between islet cells?

Answer 64

it does not require the manipulation of very many genes

Question 65

the over expression of what can make beta cells from alpha cells?

Answer 65

Pax4

Question 66

what are three ways to make functional beta cells?

Answer 66

1. directed differentiation of ESC/iPSC 2. reprogramming of islets cells/acinar cells/hepatocytes 3. inducing replication of beta cells

Question 67

what does directed differentiation try to mimic?

Answer 67

normal development

Question 68

what does directed differentiation currently involve?

Answer 68

an in vivo transplantation step

Question 69

why do we need to be careful about injected beta cells derived from directed differentiation?

Answer 69

need to be sure that they have been fully reprogrammed so that they will not give rise to cancer

Question 70

what is a downside of using alpha cells for reprogramming?

Answer 70

they do not proliferate very well and so will need to obtain lots of them

Question 71

how is the co-scientific director of the Harvard stem cell institute? and what field is he now at the forefront of?

Answer 71

professor Doug Melton | the field of generating and reprogramming beta cells

Question 72

when and what was the first evidence that beta cells could be generated in vitro? and what were they then showed to do?

Answer 72

- in 2014, Professor Doug Melton helped define a differentiation protocol for hPSC in vitro - when transplanted into diabetic mice they restored glucose regulation

Question 73

what was the second strategy for generating beta cells was Doug Melton also involved in defining?

Answer 73

generating beta cells through the reprogramming of adult exocrine cells in vivo

Question 74

how were the specific transcription factors delivered to the exocrine cells in order to reprogram them and what was the downside of this?

Answer 74

retroviral delivery | would not want to give patient cells made by this method as it might lead to cancer

Question 75

when adult exocrine cells were reprogrammed in vivo what was observed?

Answer 75

beta cells were not confined to the islets and were distributed throughout the pancreas

Question 76

what needs to be determined about location of beta cells in the pancreas?

Answer 76

whether their localisation in the islets are necessary for their function

Question 77

what did a third study do to overcome the problem of Doug Meltons second study?

Answer 77

they used small molecules facilitate the reprogramming of mouse fibroblasts into pancreatic lineage