The changes in tissue histomorphology and quality of DNA in healthy human autopsied tissues stored at -20 °C and -150 °C.

The aim of this study was to compare the changes in tissue histomorphology and DNA quality in six different healthy tissues (brain, heart, lung, liver, spleen and kidney) exempted during autopsy of healthy individuals and storage at -20 °C and -150 °C three month. Tissue samples were obtained, divided by tissue and temperature group, and for each sample, tissue histomorphology and DNA (isolated from all tissues in duplicated - 72 samples of DNA) quality were analysed. Morphology of tissue samples was studied using H&E staining. DNA was isolated using the phenol-chloroform-isoamyl alcohol method. To assess the concentration and purity of the DNA samples, we used a spectrophotometer to measure absorbance at wavelengths of 280 nm and 260 nm. The fragments of human telomerase reverse transcriptase (hTERT) gene were amplified from the DNA using PCR reaction and then visualised using the 2 % agarose gel. Samples stored at -150 °C sustained the highest degree of histomorphological damage, while samples stored at -20 °C were less degraded, compared to control. The liver samples stored at -20 °C had a mean DNA concentration (1030.4 ± 51.5 ng/µl) higher than the samples of liver tissue stored at -150 °C (497.4 ± 167.1 ng/µl) (p < 0.001). Other tissues did not have statistically significantly different DNA concentration at both temperatures. Liver samples at -20 °C had degraded DNA, showed as the absence of hTERT gene in most of samples. Other tissue samples in both temperature groups had unfragmented DNA. Storing tissue samples at -20 °C is not inferior in terms of DNA yield and integrity, and possibly superior for tissue histomorphology, comparing with samples stored at -150°C.

Clinical significance of microvessel density and proliferation in prostate cancer core biopsy.

PURPOSE: To investigate the microvessel density (MVD) and proliferation in prostate cancer (PC) core biopsies. METHODS: Core biopsy samples of PC tissue from 45 patients were routinely processed and embedded in paraffin. The samples of PC formed the investigated group (n=25), while samples of benign prostatic hyperplasia (BPH) served as controls (n=20). From paraffin blocks, 3-5 µm-thick sections were made and routine hematoxylin-eosin method and immunohistochemical ABC method with Ki67 and CD34 antibodies were applied. Immunohistochemical expression of Ki67 and CD34 was stereometrically quantified. RESULTS: The median number of Ki67 and CD34 positive cells per mm2 in PC were significantly higher in comparison to the median of these cells in BHP. The average age and Gleason score in patients with high proliferation index (proIDX) and MVD index (mvdIDX) was significantly greater in comparison to those with low proIDX and low mvdIDX. The absolute values of Ki67 expression were in highly positive and significant correlation with the absolute values of CD34 expression. Highly significant correlation was found between Gleason score and proIDX and mvdIDX. CONCLUSION: This study showed that PC expressed significantly higher values of Ki67 and CD34 in comparison to BPH. The values of proIDX and mvdIDX obtained by core biopsy could clearly show the level of cancer progression expressed through highly correlated Gleason score. In this way it is possible to identify the patients at high risk for disease progression.

Association of pulmonary histopathological findings with toxicological findings in forensic autopsies of illicit drug users.

BACKGROUND/AIM: Drug abuse remains a significant social problem in many countries. The aim of the study was to estimate association between pulmonary histopathological changes and results of toxicological analyses in forensic autopsies of illicit drug users. METHODS: This investigation was performed in the Institute of Forensic Medicine, Belgrade, and in the Clinical Center, Department of Forensic Medicine, Kragujevac, from 2000 to 2004, and included 63 medicolegal autopsies of heroin or other drug consumers who suddenly died. Autopsies, postmortem toxicological examination of drugs and serological analyses of anti-HIV/HBV/HCV antibodies were performed. RESULTS: The deceased persons were mostly male, 46/63 (73.01%), ranged in age from 19 to 49 years (mean 31 years) and all were whites. Postmortem toxicological examination was performed on all of the deceased persons and drugs in the fatal range were identified in only eight of them (12.7%), in the toxic range in ten (15.87%), and in minimal concentrations in 35 (55.56%) of the deceased persons. Drugs identified in the fatal, toxic or minimal range included heroin-morphine (38/53), cocaine (4/53), tramadol (3/53), and lorazepam (1/53). In the 7 remaining subjects, ethanol in combination with heroin was found in 4 cases, and diazepam in combination with heroin in 3 cases. Dominant pathomorphological changes were findings in the lung tissue. Most common histological changes observed in drug users were pulmonary edema--55/63 (87.3%), acute alveolar hemorrhages--49/63 (77.78%), hemosiderin-laden macrophages (siderophages)--52/63 (82,54%), and emphysematous changes--51/63 (80,95%). CONCLUSION: Pulmonary edema is the frequent non-specific autopsy finding which is associated with virtually all routes of drug administration. The histopatological study is necessary to determinate a cause of death when a deceased person has the history of dependence or abouse of psychoactive drugs with negative toxicological results.