Alzheimer’s disease

Alzheimer’s disease : 

Alzheimer’s disease By Mohammed Razeeth s

Phosphoylation of tau : 

Phosphoylation of tau In AD tau protein get hyperphosphorylated due to Mutation Activities of kinase Regulation of protein phosphtase Quinolinic Acid

Serine and threonine residues : 

Serine and threonine residues To date, at least 37 serine and threonine residues have been found to be phosphorylated in PHF-tau Thr39, Ser46, Thr69,Thr123, Ser137, Thr153, Thr175, Thr181, Ser198, Ser199,Ser202, Thr205, Ser208, Ser210, Thr212, Ser214, Thr217,Thr231, Ser235, Ser237, Ser238, Ser241, Ser262, Ser285,Ser305, Ser324, Ser352, Ser356, Ser396, Ser400, Thr403,Ser404, Ser409, Ser412, Ser413, Ser416, and Ser422

Serine and threonine residues : 

Serine and threonine residues Some of the phosphorylation sites seen in PHF tau are not phosphorylated at all in normal brains. These sites include Thr212/Ser214, Thr231/Ser235 [90], and Ser422[91, 92].

Phosphoryltion at Tyr : 

Phosphoryltion at Tyr The phosphorylation site of tau was mapped as Tyr18. Williamson et al [94] demonstrated that in primary human and rat brain cortical cultures tau is phosphorylated at Tyr 29 upon treatment with Aβ

Regulation : 

Regulation

MAP : 

MAP Hyper phosphorylation tau gain a toxic activity to sequester normal tau and other MAPs, such as MAP1 and MAP2, and cause microtubule disassembly

Kinase involve in phospho.. : 

Kinase involve in phospho.. GSK-3β, cdk5, cAMP-dependent protein kinase (PKA), stress-activated protein kinases, and calcium/calmodulin-dependent kinase II (CaMK-II)

Pathway : 

Pathway

Evidance : 

Evidance

My concern : 

My concern To up regulate protein phosphates by activating PP2A

Evidance : 

Evidance

Methodology : 

Methodology Cell lineHT22 hippocampal neuronal cell line, SH-SY5Y Retroviral infection Immuno chemical method-immuno histo compatability SDS page

Retro viral infection : 

Retro viral infection For single round of infection Add 4 μg/ml Polybrene to virus (from 100 x stock), remove medium and cover cells with virus + Polybrene

Slide 16: 

12-well plate: 300-500 μl/well 6-well plate: 750-1000 μl/well T25/5 cm dish: 1.5 ml 10 cm dish: 4.5 ml

Day 1, 6-8 hrs later : 

Day 1, 6-8 hrs later Add medium (the type in which the target cells grow) to the virus incubations to dilute the Polybrene which is toxic at concentrations higher than 2 μg/ml. Eg; 500 μl of virus, add 750 μl of medium 1 ml of virus, add 1.5 ml medium

For 2 rounds of infection : 

For 2 rounds of infection 1. Start with the first infection on the morning of Day 1 as described above In the morning of Day 2, remove the virus containing medium Leave the cells for 24 hrs, until Day 3.

For 3 rounds of infection : 

For 3 rounds of infection Start with the first infection in the late afternoon/evening of Day 1 by removing medium from target cells and adding virus + Polybrene, leave overnight, do not dilute polybrene.

For 3 rounds of infection : 

For 3 rounds of infection The next morning remove virus and replace by new virus + Polybrene. In evening do not remove virus of second infection, but dilute it by adding an equal amount of new virus without polybrene.

For 4 rounds of infection : 

For 4 rounds of infection In the morning of Day 2, remove the virus containing medium of the first infection and repeat the infection as described above. In the evening of Day 2, remove the virus containing medium of the first infection and repeat the infection. In the morning of Day 3 repeat the infection and discard the Phoenix cells.

For 4 rounds of infection : 

For 4 rounds of infection In the evening of Day 3, change the medium of the infected cells. 24 hours later, start antibiotic selection. For puromyocin, select for 3 days. For hygromyocin, select for 7 days.

Slide 24: 

Thank you