Attenuated DNA damage repair by trichostatin A through BRCA1 suppression

Recent studies have demonstrated that some histone deacetylase (HDAC) inhibitors enhance cellular radiation sensitivity. However, the underlying mechanism for such a radiosensitizing effect remains unexplored. Here we show evidence that treatment with the HDAC inhibitor trichostatin A (TSA) impairs radiation-induced repair of DNA damage. The effect of TSA on the kinetics of DNA damage repair was measured by performing the comet assay and gamma-H2AX focus analysis in radioresistant human squamous carcinoma cells (SQ-20B). TSA exposure increased the amount of radiation-induced DNA damage and slowed the repair kinetics. Gene expression profiling also revealed that a majority of the genes that control cell cycle, DNA replication and damage repair processes were down-regulated after TSA exposure, including BRCA1. The involvement of BRCA1 was further demonstrated by expressing ectopic wild-type BRCA1 in a BRCA1 null cell line (HCC-1937). TSA treatment enhanced radiation sensitivity of HCC-1937/wtBRCA1 clonal cells, which restored cellular radiosensitivity (D(0) = 1.63 Gy), to the control level (D(0) = 1.03 Gy). However, TSA had no effect on the level of radiosensitivity of BRCA1 null cells. Our data demonstrate for the first time that TSA treatment modulates the radiation-induced DNA damage repair process, in part by suppressing BRCA1 gene expression, suggesting that BRCA1 is one of molecular targets of TSA.     

Modeling of Hepatocytes Proliferation Isolated from Proximal and Distal Zones from Human Hepatocellular Carcinoma Lesion

Isolation of hepatocytes from cirrhotic human livers and subsequent primary culture are important new tools for laboratory research and cell-based therapeutics in the study of hepatocellular carcinoma (HCC). Using such techniques, we have previously identified different subpopulations of human hepatocytes and among them one is showing a progressive transformation of hepatocytes in HCC-like cells. We have hypothesized that increasing the distance from the neoplastic lesion might affect hepatocyte function and transformation capacity. However, limited information is available in comparing the growth and proliferation of human hepatocytes obtained from different areas of the same cirrhotic liver in relation to their distance from the HCC lesion. In this study, hepatocytes from 10 patients with cirrhosis and HCC undergoing surgical resections from specimens obtained at a proximal (CP) and distal (CD) distance from the HCC lesion were isolated and placed in primary culture. CP hepatocytes (CP-Hep) were isolated between 1 to 3 cm (leaving at least 1cm margin to avoid cancer cells and/or satellite lesions), while CD hepatocytes (CD-Hep) were isolated from more than 5 cm or from the contralateral-lobe. A statistical model was built to analyze the proliferation rates of these cells and we evaluated expression of HCC markers (Glypican-3 (GPC3), αSmooth Muscle Actin (α-SMA) and PCNA). We observed a significant difference in proliferation and in-vitro growth showing that CP-Hep had a proliferation pattern and rate significantly different than CD-Hep. Based on these data, this model can provide information to predict growth of human hepatocytes in primary culture in relation to their pre-cancerous state with significant differences in the HCC markers expression. This model provides an important innovative tool for in-vitro analysis of HCC.     

Deferiprone: structural and functional modulating agent of hemoglobin fructation

Diabetic complication arises from the presence of advanced glycation end products in different sites of the body. Great attention should be paid to recognizing anti-glycation compounds. Here, deferiprone as an oral iron chelator drug administrated in treatment of β-thalassemic patients was selected to find its effect on the fructation of hemoglobin (Hb). Our results indicated that deferiprone could prevent the AGE and carbonyl formation via inhibition of structural changes in the structure of Hb during the fructation process. Moreover, deferiprone can preserve peroxidase and esterase activities of fructated Hb similar to native Hb. Therefore, deferiprone can be introduced as an anti-glycation drug to prevent the AGE formation.     

Evidence for crossing the blood barrier of adult rat brain by human adipose-derived mesenchymal stromal cells during a 6-month period of post-transplantation

Background aimsTherapeutic promises of adult stem cells have been overshadowed by an elicited immune response, low maintenance of implanted cells or concerns regarding their migration to non-target sites. These problems might be lessened by the use of immune privilege cells and tissues for implantation.MethodsIn this study, human adipose-derived mesenchymal stromal cells (hADMSCs) were stably transfected with a vector containing Turbo green fluorescent protein (GFP) and JRed, which allows tracing the cells after transplantation. Labeled hADMSCs were transplanted into the adult rat brain followed by assessment of their survival and migration during 6 months after transplantation.ResultsResults indicate that there were no postsurgical complications, and the animals thrived after transplantation. The lesions of the surgical process were remarkable at the first weeks, and a high number of transplanted cells were accumulated around them. Cell populations declined over time as they partly migrated away from the injection sites; nonetheless, they were detectable at each examination time point. Although the cells could survive and remain at the injection site for up to 6 months, some of them drifted to spleen, which is an indication of their ability to cross the blood-brain barrier.ConclusionsDespite the high survival rate of hADMSCs in the xenogenic condition, which is an ideal criterion in cell therapy, irregular migration tendency must be handled with caution.     

Incidence of first ever stroke during Hajj ceremony

Background

The Hajj Ceremony, the largest annual gathering in the world, is the most important life event for any Muslim. This study was designed to evaluate the incidence of stroke among Iranian pilgrims during the Hajj ceremony.

Methods

We ascertained all cases of stroke occurring in a population of 92,974 Iranian pilgrims between November 27, 2007 and January 12, 2008. Incidence and risk factors of the first ever stroke in Hajj pilgrims were compared, within the same time frame, to those of the Mashhad residents, the second largest city in Iran. Data for the latter group were extracted from the Mashhad Stroke Incidence Study (MSIS) database.

Results

During the study period, 17 first-ever strokes occurred in the Hajj pilgrims and 40 first-ever stroke strokes occurred in the MSIS group. Overall, the adjusted incidence rate of first ever stroke in the Hajj cohort was lower than that of the MSIS population (9 vs. 16 per 100,000). For age- and gender-specific subgroups, the Hajj stroke crude rates were in general similar to or lower than the general population of Mashhad, Iran, with the exception of women aged 35 to 44 years and aged >75 years who were at greater risk of having first-ever stroke than the non-pilgrims of the same age.

Conclusion

The first ever stroke rate among Iranian Hajj pilgrims was lower than that of the general population in Mashhad, Iran, except for females 35–44 or more than 75 years old. The number of events occurring during the Hajj suggests that Islamic countries should consider designing preventive and screening programs for pilgrims.

     

Isolation and characterization of a novel scFv antibody fragments specific for Hsp70 as a tumor biomarker

Many studies have shown that more than 50% of tumors express heat shock protein 70 kDa (Hsp70) at the plasma membrane surface while not seen in normal cells, therefore it is a promising therapeutic target in human cancers. Hence, we used phage display technology to produce a single-chain fragment variable (scFv) antibody against human Hsp70. For this, a target peptide from human Hsp70 was designed using bioinformatics studies and was chemically synthesized. Then, the selection was performed using four rounds of biopanning with a stepwise decreased amount of the target peptide. Fourteen positive scFv clones were selected using monoclonal phage enzyme-linked immunosorbent assay screening, which was further characterized by means of the polymerase chain reaction and DNA sequencing. Among them, the G6 clone was selected to express scFv into the Escherichia coli. Expression and purification of the scFv shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and confirmed by Western blot analysis. In silico analysis confirmed specific binding of the scFv to Hsp70 in CDR regions. The specificity of the scFv measured by surface plasmon resonance and immunofluorescence of the A549 human lung carcinoma cell line confirmed the in vitro function of the scFv. Based upon these findings, we propose a novel anti-human Hsp70 scFv as potential immunotherapy agents that may be translated into preclinical/clinical applications.