Investigation of Optimum pH and Temperature for In-Vitro Crystallization of Urinary Cystine

Cystinuria contributes in formation of urinary stones. But, it has been reported that cystinuria is diagnosed when someone experiences with cystine stones. Therefore, early diagnosis of this condition is important. Thus, the objective of the study was to determine the optimum pH and temperature for crystallization of urine cystine in-vitro. Cystinuria solutions were prepared with the concentrations of 40, 60, 70, 75, 80, 90, 100 and 120 mg/dL. The pH of each solution was changed with the addition of acetic acid. Then solutions were exposed to temperature +4°C and 37°C, for 15, 30 and 45min. The sediments were observed microscopically for cystine crystals formation. Then acetone was added to cystinuria with the ratio of cystinuria:acetone, 8:1, 4:1, 2:1 and 1.1 and pH was altered with acetic acid and were subjected to +4 °C and 37 °C, for 15, 30 and 45 minutes and sediment was observed for cystine crystals under the microscope. Cystine crystallization had been occurred in the cystinuria of ≥100 mg/dL at pH 5 at 37 ° C and +4 °C, 30min after the addition of acetic acid whereas with the addition of acetone at cystinuria of ≥75mg/dL at pH 5 in both 37°C and at +4°C, 30min after the addition of acetic acid. The number of cystine crystals per High Power Field (HPF) was highest where cystinuria:acetone was 8:1.  The optimum conditions for cystine crystallization is at pH 5, 37 °C and +4 °C, 30min after acidifying with acetic acid at the minimum concentration of 100 mg/dL  of cystinuria. With the addition of acetone, at the ratio of cystinuria:acetone 8:1 with minimum concentration of cystinuria of 75 mg/dL.   KEYWORDS: Cystine, Crystallization, Acetic acid, Acetone, Temperature, p

Occurrence of Urinary Crystals among Urinary Tract Infections Suspected Paediatric Patients, Sri Lanka

Crystalluria has become one of the most vital biomarkers in urinalysis in detecting several disease conditions. It has been reported that urinary tract infections (UTI) may be the presenting sign of Urolithiasis in children. Therefore, the objective of this study was to identify and estimate the different types of crystals in the urine samples collected from UTI suspected children who admitted to the Lady Ridgeway Hospital for children, Sri Lanka. A descriptive cross-sectional study was conducted using 400 children belong to age<12 years suspected with UTI. The participants included 242 males and 158 females. The urine samples were collected prior to start antibiotics. Each sample was examined macroscopically and centrifuged at 2000 rpm for 5 minutes. The urine sediment was examined under the light microscope and different crystal types were identified and counted at x40 magnification. Out of 400 samples 82 samples (82/400) were positive for crystalluria. The crystal types present were uric acid, calcium oxalate, triple phosphate, ammonium biuate and ammonium urate. None of the samples showed abnormal crystal types. The distribution of each crystal type was as follow; uric acid 25/82, calcium oxalate 34/82, triple phosphate 12/82, ammonium biuate 7/82 and ammonium urate 4/82. The quantity of crystals per mL of urine was ranged as follow; uric acid 850-130,000, calcium oxalate 350- >250,000, triple phosphate 650-6,000, ammonium biurate and ammonium urate were presented in clumps. KEYWORDS: Crystalluria, Uric Acid, Calcium Oxalate, Triple Phosphate, Ammonium Biurate, Ammonium Urate, Urolithiasis, Urinary Tract Infections

Bioinformatic identification of proteins with tissue-specific expression for biomarker discovery

<p>Abstract</p> <p>Background</p> <p>There is an important need for the identification of novel serological biomarkers for the early detection of cancer. Current biomarkers suffer from a lack of tissue specificity, rendering them vulnerable to non-disease-specific increases. The present study details a strategy to rapidly identify tissue-specific proteins using bioinformatics.</p> <p>Methods</p> <p>Previous studies have focused on either gene or protein expression databases for the identification of candidates. We developed a strategy that mines six publicly available gene and protein databases for tissue-specific proteins, selects proteins likely to enter the circulation, and integrates proteomic datasets enriched for the cancer secretome to prioritize candidates for further verification and validation studies.</p> <p>Results</p> <p>Using colon, lung, pancreatic and prostate cancer as case examples, we identified 48 candidate tissue-specific biomarkers, of which 14 have been previously studied as biomarkers of cancer or benign disease. Twenty-six candidate biomarkers for these four cancer types are proposed.</p> <p>Conclusions</p> <p>We present a novel strategy using bioinformatics to identify tissue-specific proteins that are potential cancer serum biomarkers. Investigation of the 26 candidates in disease states of the organs is warranted.</p

A secretome profile indicative of oleate-induced proliferation of HepG2 hepatocellular carcinoma cells

Increased fatty acid (FA) is often observed in highly proliferative tumors. FAs have been shown to modulate the secretion of proteins from tumor cells, contributing to tumor survival. However, the secreted factors affected by FA have not been systematically explored. Here, we found that treatment of oleate, a monounsaturated omega-9 FA, promoted the proliferation of HepG2 cells. To examine the secreted factors associated with oleate-induced cell proliferation, we performed a comprehensive secretome profiling of oleate-treated and untreated HepG2 cells. A comparison of the secretomes identified 349 differentially secreted proteins (DSPs; 145 upregulated and 192 downregulated) in oleate-treated samples, compared to untreated samples. The functional enrichment and network analyses of the DSPs revealed that the 145 upregulated secreted proteins by oleate treatment were mainly associated with cell proliferation-related processes, such as lipid metabolism, inflammatory response, and ER stress. Based on the network models of the DSPs, we selected six DSPs (MIF, THBS1, PDIA3, APOA1, FASN, and EEF2) that can represent such processes related to cell proliferation. Thus, our results provided a secretome profile indicative of an oleate-induced proliferation of HepG2 cell

Low-Dose Cd Induces Hepatic Gene Hypermethylation, along with the Persistent Reduction of Cell Death and Increase of Cell Proliferation in Rats and Mice

Cadmium (Cd) is classified as a human carcinogen probably associated with epigenetic changes. DNA methylation is one of epigenetic mechanisms by which cells control gene expression. Therefore, the present study genome-widely screened the methylation-altered genes in the liver of rats previously exposed to low-dose Cd.Rats were exposed to Cd at 20 nmol/kg every other day for 4 weeks and gene methylation was analyzed at the 48(th) week with methylated DNA immunoprecipitation-CpG island microarray. Among the 1629 altered genes, there were 675 genes whose promoter CpG islands (CGIs) were hypermethylated, 899 genes whose promoter CGIs were hypomethylated, and 55 genes whose promoter CGIs were mixed with hyper- and hypo-methylation. Caspase-8 gene promoter CGIs and TNF gene promoter CGIs were hypermethylated and hypomethylated, respectively, along with a low apoptosis rate in Cd-treated rat livers. To link the aberrant methylation of caspase-8 and TNF genes to the low apoptosis induced by low-dose Cd, mice were given chronic exposure to low-dose Cd with and without methylation inhibitor (5-aza-2'-deoxyctidene, 5-aza). At the 48(th) week after Cd exposure, livers from Cd-treated mice displayed the increased caspase-8 CGI methylation and decreased caspase-8 protein expression, along with significant increases in cell proliferation and overexpression of TGF-β1 and cytokeratin 8/18 (the latter is a new marker of mouse liver preneoplastic lesions), all which were prevented by 5-aza treatment.These results suggest that Cd-induced global gene hypermethylation, most likely caspase-8 gene promoter hypermethylation that down-regulated its expression, leading to the decreased hepatic apoptosis and increased preneoplastic lesions