Biomedical subjects

Hari Koul

Publications and source records attributed to Hari Koul.

4 recordsLinked to original sources

PubMed2006

Global gene expression patterns in mouse wolffian duct development.

PURPOSE: We identified genes responsible for terminal differentiation of the mouse ureter. MATERIALS AND METHODS: We isolated the wolffian ducts of Black Swiss mice from embryonic days 12.5 to 14.5. These ducts were processed for total RNA extraction. RNA was amplified and converted to biotinylated cDNA, which was hybridized to Affymetrix(R) mouse 430 microarray GeneChips. Data were analyzed using Affymetrix software provided by the core facility at our institution. Data were confirmed by semiquantitative polymerase chain reaction and immunohistochemistry. RESULTS: Of 42,000 evaluable transcripts 412 were up-regulated and 133 were down-regulated more than 1.5-fold between embryonic days 12.5 and 14.5. Of up-regulated genes 17 were up-regulated more than 3-fold. Of these genes 5 were selected for further study and they were identified as having roles in cell growth/regulation in general. Increased expression of Foxa1 was seen at embryonic day 14.5 in the distal ureter and urogenital sinus. CONCLUSIONS: To our knowledge this is the first report of differential gene expression patterns in the developing mouse wolffian duct. Our results diverge from expression patterns reported in the kidney. This information will enable future comparisons between WT and transgenic mice with an abnormal phenotype.

Animals↗

PubMed2005

Cellular transfection to deliver alanine-glyoxylate aminotransferase to hepatocytes: a rational gene therapy for primary hyperoxaluria-1 (PH-1).

BACKGROUND: Primary hyperoxaluria-type 1 (PH-1) is a rare autosomal recessive disorder of glyoxalate metabolism caused by deficiency in the liver-specific peroxisomal enzyme alanine-glyoxalate transaminase 1 (AGT) resulting in the increased oxidation of glyoxalate to oxalate. Accumulation of oxalate in the kidney and other soft tissues results in loss of renal function and significant morbidity. The present treatment options offer some relief in the short term, but they are not completely successful. In the present study, we tested the feasibility of corrective gene therapy for this metabolic disorder. METHODS: A cDNA library was made from HepG2 cells. PCR primers were designed for the AGT sequence with modifications to preclude mistargeting during gene delivery. Amplified AGT cDNA was cloned as a fusion protein with green fluorescent protein (GFP) using the vector EGFP-C1 (Clontech) for monitoring subcellular distribution. Sequence and expression of the fusion protein was verified. Fusion protein vectors were transfected into hepatocytes by liposomal transfection. AGT expression and subcellular distribution was monitored by GFP fluorescence. RESULTS: HepG2 cells express full-length mRNA coding for AGT as confirmed by insert size as well as sequence determination. Selective primers allowed us to generate a modified recombinant GFP-AGT fusion protein. Cellular transfections with Lipofectamine resulted in transfection efficiencies of 60-90%. The recombinant AGT did localize to peroxisomes as monitored by GFP fluorescence. CONCLUSIONS: The results demonstrate preliminary in vitro feasibility data for AGT transfection into the hepatocytes. To the best of our knowledge, this is the first study to attempt recombinant AGT gene therapy for treatment of primary hyperoxaluria-1.

Cells, Cultured↗

PubMed2005

The molecular basis of erectile dysfunction: from bench to bedside.

Erectile dysfunction is a common problem affecting many men across all age groups. Its etiology is multifactorial. Hormonal, vascular, neurogenic, lifestyle, and psychological entities have all been implicated as causative agents. The molecular basis underlying its etiology and progression is complex and still challenges researchers in the field. Nonetheless, newly discovered common pathways and targets of its pathogenesis have opened a new era for both prevention and active treatment of the disease. This review describes some of the known molecular mechanisms contributing to erectile dysfunction and discusses the future of gene therapy for the disease.

Journal Article↗

PubMed2003

Oxalate exposure promotes reinitiation of the DNA synthesis and apoptosis of HK-2 cells, a line of human renal epithelial cells.

Oxalate, a metabolic end product, is a major constituent of kidney stones. Previously, we and others have demonstrated that oxalate is toxic to renal epithelial cells. In the present study, we characterized oxalate-induced cell death in HK2 cells, a line of renal epithelial cells from the human kidney. For these studies, HK2 cells were exposed to oxalate for various time points. Cells were examined for nuclear morphology, DNA fragmentation, and expression of various apoptosis-related proteins. Apoptotic mode of cell death was also confirmed by TUNEL assay. Results from these studies revealed that oxalate exposure resulted in time-dependent increase in DNA fragmentation. Maximum DNA fragmentation was observed at 2-24 hours following oxalate exposure. Results from Western blot analysis demonstrated an increased expression of the FAS ligand. Taken together, these data reveal that oxalate-associated nephrotoxicity results from oxalate-induced apoptosis of renal epithelial cells.

Apoptosis↗