clinical biochemistry

which diabetes is insulin dependant?

Type 1 diabetes

which diabetes is insulin resistant?

Type 2 diabetes

Hyperglycaemia

blood glucose to high

hypoglycaemia

blood glucose to low

Fed state -> food broken down…

glucose levels rise-> insulin released

where is glucose stored?

• skeletal muscle fat tissue/ adipose tissue

Glycolysis

glucose -> stores

Starved state -> glucagon …

signals release of glucose

Gluconeogenesis

stores -> glucose

Insulin produced where?

Beta cells in islets of Langerhans in pancreas

Normally Increase in blood glucose levels

GLUT2 transporters allow glucose into beta cells -> Beta cells release insulin

Beta cells transporters

GLUT2 transporters

Normally with no insulin adipose / skeletal muscles

can’t uptake glucose

Once insulin released

insulin binds to the surface of skeletal muscle/adipose tissue cells

Transporter on adipose/skeletal muscle tissue cells

GLUT4 transporter allows glucose to enter those cells

Insulin and liver

insulin signals liver to stop producing glucose-maintaining homeostatic level

With diabetes type 1 -> insulin…

not produced at all

What is causing hyperglycaemia with type 1 diabetes?

glucose is not taken up for energy nor storage. Liver keeps producing glucose (gluconeogenesis). Glucose from food aswell.

With type 2 diabetes -> insulin…

made by beta cells and respond to fed state. When binding to adipose/ skeletal muscle tissue cells - the signal is weak so more insulin is required to meet requirements.

Why do beta cells die with type 2 diabetes?

the get stressed from producing too much insulin to meet required signal for sink cells

Which type of diabetes is autoimmune?

Type 1

Which type of diabetes is an acquired metabolic disorder?

Type 2

In Type 1 diabetes (2)

1. viewed as foreign: Insulin and GADA 2. Islets of Langerhans destroyed

In Type 2 diabetes (2)

1. Insulin produced inefficiently 2. Beta cells over work stress die.

Idiopathic diabetes

we don’t know the cause.

Glucosuria

glucose in urine

Polyuria

frequent urination

Polydipsia

thirst

With diabetes in reference with kidneys

not all glucose is reabsorbed -creating osmotic pressure- less water reabsorbed- glucose in urine

Renal threshold ( when kidneys let glucose pass through urine)

11mmol/L

With diabetes- overall change in levels (3)

1. Decreased glucose uptake

2. Increased glycogenolysis

3. Increased gluconeogenesis

How does the liver make glucose during a starved state?

1. stored fat -> FFA and glycerol (lipolysis)

2. Transported in bloodstream to liver.

3. FFA oxidised to Acetyl CoA

4. Acetyl CoA and glycerol drives gluconeogenesis

5. Increase in blood glucose

How are ketones produced?

Acetyl CoA -> ketones in the liver

Diabetes and ketones

insulin inhibits acetyl CoA -> ketones

Insulin deficiency increases(diabetes and ketones):

• FFA supply to liver and therefore Acetyl CoA -Ketone production - Elevated serum ketones

Hyperventilation

lower blood pH

Diabetic Ketoacidosis (4):

1. Ketones are acidic-lowering Blood pH due to carboxyl gp.

2. Hyperventilation

3. Respiration increases & exhale CO2

4. Nausea & vomiting & dehydration

Hyperglycaemia problems over time

glucose + proteins -> glycated proteins (leads to cell & tissue dysfunction & blood vessel damage)

Long term Hyperglycaemia increases risk of (4):

1.retinopathy

2. Nephropathy

3. Neuropathy

4. Cardiovascular disease

Diagnosing Diabetes (3):

1. urine dipstick - checks for blood/glucose/ketone/pH/proteins

2. Fasting blood glucose test - measures baseline blood glucose

3. Oral glucose tolerance test - indicates ability of beta cells to secrete insulin

Biomarker for blood glucose

HbA1c - allows longterm estimation of blood glucose

testing for biomarker (3)

1. take sample

2. chemical and biochemical analysis to detect biomarkers

3. compare to references

plasma vs. serum

serum has clotting factors,

plasma has been treated with anticoagulants

• serum = plasma - fibrinogen

Blue cap

reversible inhibition of clotting

clotting tests

yellow cap

allows clots,

serum for analysis of soluble components

pink/ purple cap

irreversible inhibition of clotting

blood cell/ plts analysis

green cap

anticoagulant

hormone/chromosome analysis

Grey cap

halts metabolism

analysis of energy sources, glucose

diagnostic biomarkers

they detect disease/ conditions

monitoring biomarkers

to assess status of disease

prognostic biomarkers

they predict disease progression

risk biomarkers

to see the transition from healthy to diseased

how do biomarkers levels change with extensive cell damage?

intracellular molecules (cell type specific) released

how do biomarkers levels change with cell proliferation?

increased markers of cell growth / rapid proliferation produced by cells

how do biomarkers levels change with enzyme induction?

increased expression of cell specific membranes

how do biomarkers levels change with duct obstruction?

blocked flow of molecules

increased in blood

spectroscopy with analysis of markers

colorimetric assays, chemical rxns, increased enzyme activity

chromatography with analysis of markers

molecule separation based on size

mass spectrometry with analysis of markers

molecule identification based on mass

immunoassays with analysis of markers

molecule identification based on molecular sequence/strructure

detected w/ antibodies

molecular techniques with analysis of markers

DNA/RNA sequencing to identify infection and genetic markers

POCT

point of care testing

patient present

rapid turnaround

non-laboratory

Clinical Biochemistry Department

labs

greater range of test

automated

Good Biomarkers should have:

• specificity

• biological understanding

• reproducibility

• efficient

• cost effective

• feasible sample collection

• better than gold standard

• sensitivity

pre- analytical factors affecting test result

• technical - correct patient and correct labelling of sample

• biological - age, gender, mass, ethnicity

analytical factors affecting test result

accuracy and precision

post analytical factors affecting test result

reporting, transmission, transcript, clinical decision

how is reference range calculated

$\mu$ ± 2SD

Liver Functions (4)

• blood proteins production- albumin, clotting factors

• bile production

• bilirubin excretion- haem breakdown

• phagocytosis of bacteria- via kupffer cells

metabolism and detox in the liver

• glycogen → glucose

• fatty acid metabolism -ketone production

• a.a. metabolism - ammonia→ urea

pre-hepatic with bilirubin as a biomarker

• high serum of unconjugated bilirubin

• other LFT normal

• eg. haemolytic disease

hepatic with bilirubin as a biomarker

• high serum of conjugated bilirubin

• liver enzymes elevated, liver damage

• eg. hepatitis

post-hepatic with bilirubin as a biomarker

• high serum of conjugated bilirubin

• blockage of bile flow-cholestasis

• eg. gallstones

LFT meaning

liver function test

elevated unconjugated bilirubin means…

• increased bilirubin synthesis

• haemolytic disease

• decreased conjugation by liver → genetic disease

• neonatal jaundice

elevated conjugated bilirubin means…

• hepatocellular disease → liver damage

• cholestasis → blockage to flow of bile

• decreased transport of conjugated bilirubin

acute liver disease is…

viral, toxic, due to alcohol consumption

liver inflammation

rapid onset

chronic liver disease is …

alcoholic hepatitis/cirrhosis, non-alcoholic steatohepatitis, autoimmune disorders, genetics

slow progression

causes of acute hepatitis

hep A, hep B, Hep C (viral)

Paracetamol (toxins)

causes of chronic hepatitis

ongoing hepatic necrosis & liver inflammation

ongoing over consumption of alcohol, some medications, recreational drugs

acute and chronic hepatitis change levels of:

• increased ALT (alanine aminotransferase)

• increased AST (aspartate aminotransferase)

• increased conjugated bilirubin

what is cholestasis

a blockage in the bile duct, meaning bile flow is obstructed into duodenum

results in bile (w/ conjugated bilirubin) is absorbed into the blood →obstructive jaundice

intrahepatic cholestasis

• structural liver tissue damage

• decreased bile flow in absence of overt bile duct obstruction

• accumulation of bile constituents in liver and blood

extrahepatic cholestasis

• bile duct obstruction, eg. gallstones, malignancies, pancreatitis

• increased ALP

• increased conjugated bilirubin

Alkaline Phosphatase (ALP) found where

• surface of hepatocyte

• microvilli of bile canaliculi

• ducts

Alkaline phosphatase (ALP) why would it be raised?

• cholestasis

• hepatic injury

• infiltrative diseases, w/ tumours

• growth spurts in adolescents

• broken bones