Availability of individual proteins for quantitative analysis in postmortem brains preserved in two different brain banks.

AIM: Postmortem brain research is necessary for elucidating the pathology of schizophrenia; an increasing number of studies require a combination of suitable tissue samples preserved at multiple brain banks. In this study, we examined whether a comparative study of protein expression levels can be conducted using postmortem brain samples preserved in different facilities. METHODS: We compared the demographic factors of postmortem brain samples preserved in two institutions and measured and compared the expression levels of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and glial fibrillary acidic protein (GFAP) in the prefrontal cortex and superior temporal gyrus. GAPDH is generally used as a loading control for western blotting, and GFAP is considered as an astrocyte marker in the brain. RESULTS: We found significant differences between the two institutions in postmortem interval, age at death, and preservation time. To reduce the effects of these differences on our measurements, the parameters were set as covariates in our analyses of covariance. Subsequently, no differences in GAPDH and GFAP expression were found between institutions. CONCLUSIONS: When studies are conducted using brain samples preserved in different brain banks, differences in demographic factors should be carefully considered and taken into account by statistical methods to minimize their impact as much as possible. Since there was no significant difference in the protein expression levels of GAPDH and GFAP in either region between the two institutions that preserved the postmortem brains, we concluded that it is possible to perform protein quantitative analysis assuming that there is no effect of difference between two institutions.

The influence of tissue pH and RNA integrity number on gene expression of human postmortem brain.

Background: Evaluating and controlling confounders are necessary when investigating molecular pathogenesis using human postmortem brain tissue. Particularly, tissue pH and RNA integrity number (RIN) are valuable indicators for controlling confounders. However, the influences of these indicators on the expression of each gene in postmortem brain have not been fully investigated. Therefore, we aimed to assess these effects on gene expressions of human brain samples. Methods: We isolated total RNA from occipital lobes of 13 patients with schizophrenia and measured the RIN and tissue pH. Gene expression was analyzed and gene sets affected by tissue pH and RIN were identified. Moreover, we examined the functions of these genes by enrichment analysis and upstream regulator analysis. Results: We identified 2,043 genes (24.7%) whose expressions were highly correlated with pH; 3,004 genes (36.3%) whose expressions were highly correlated with RIN; and 1,293 genes (15.6%) whose expressions were highly correlated with both pH and RIN. Genes commonly affected by tissue pH and RIN were highly associated with energy production and the immune system. In addition, genes uniquely affected by tissue pH were highly associated with the cell cycle, whereas those uniquely affected by RIN were highly associated with RNA processing. Conclusion: The current study elucidated the influence of pH and RIN on gene expression profiling and identified gene sets whose expressions were affected by tissue pH or RIN. These findings would be helpful in the control of confounders for future postmortem brain studies.

Synthesis of omega-3 long-chain polyunsaturated fatty acid-rich triacylglycerols in an endemic goby, Gymnogobius isaza, from Lake Biwa, Japan.

It is commonly observed that freshwater fish contain lower amounts of omega-3 long-chain polyunsaturated fatty acids (LC-PUFAs), such as eicosapentaenoic acid (EPA, C20:5n-3) and docosahexaenoic acid (DHA, C22:6n-3), than marine fish species. In this study, we performed a detailed comparative analysis of phospholipids (PLs) and triacylglycerols (TAGs) from Gymnogobius isaza, a freshwater goby endemic to Lake Biwa inhabiting the lake bottom, and Gymnogobius urotaenia, a related goby that inhabits the shore of Lake Biwa. We found that tissues from G. isaza contain remarkably high amounts of omega-3 LC-PUFAs in both PLs and TAGs. Mass spectrometry analysis of TAGs demonstrated that the most abundant TAG molecular species were TAG (16:0/18:1/20:5), followed by TAG (14:0/18:1/20:5), in which EPA is incorporated into TAG at either the sn-1 or sn-3 positions. We isolated cDNAs encoding acyl-CoA: diacylglycerol acyltransferase designated as GiDGAT1 and GiDGAT2, from G. isaza. Expression studies using a neutral lipid-deficient Saccharomyces cerevisiae mutant strain demonstrated that both GiDGAT1 and GiDGAT2 possessed diacylglycerol acyltransferase activity, and preferential incorporation of LC-PUFA into TAG was observed in the presence of GiDGAT1. This study revealed the novel lipid profiles of G. isaza and identified the enzymes that were involved in the production of PUFA-containing TAGs.