Chartchalerm Isarankura-Na-Ayudhya

Osteoporosis is frequently found in chronic diabetic patients, and it results in an increased risk of bone fractures occurring. The underlying mechanism of osteoporosis in diabetic patients is still largely unknown. Annexin A2 (ANXA2), a family of calcium-binding proteins, has been reported to be involved in many biological process including bone remodeling. This study aimed to investigate the role of ANXA2 in mesenchymal stem cells (MSCs) during in vitro osteoinduction under high-glucose concentrations. Osteogenic gene expression, calcium deposition, and cellular senescence were determined. The high-glucose conditions reduced the osteogenic differentiation potential of the MSCs along with the lower expression of ANXA2. Moreover, the high-glucose conditions increased the cellular senescence of the MSCs as determined by senescence-associated β-galactosidase staining and the expression of p16, p21, and p53 genes. The addition of recombinant ANXA2 could recover the glucose-induced deterioration of the osteogenic differentiation of the MSCs and ameliorate the glucose-induced cellular senescence of the MSCs. A Western blot analysis revealed an increase in p53 and phosphorylated p53 (Ser 15), which was decreased by recombinant ANXA2 in MSC osteoblastic differentiation under high-glucose conditions. Our study suggested that the alteration of ANXA2 in high-glucose conditions may be one of the plausible factors in the deterioration of bones in diabetic patients by triggering cellular senescence.

Nowadays, a lot of produce (fruits and vegetables) sold in many countries are contaminated with pesticide residues, which cause severe effects on consumer health, such as cancer and neurological disorders. Therefore, this study aims to determine whether cooking processes can reduce the pesticide residues in commonly consumed vegetables (Chinese kale and yard long beans) in Thailand. For cooking experiments, the two vegetables were cooked using three different processes: boiling, blanching, and stir-frying. After the treatments, all cooked and control samples were subjected to extraction and GC-MS/MS analysis for 88 pesticides. The results demonstrated that pesticide residues were reduced by 18–71% after boiling, 36–100% after blanching, and 25–60% after stir-frying for Chinese kale. For yard long beans, pesticide residues were reduced by 38–100% after boiling, 27–28% after blanching, and 35–63% after stir-frying. Therefore, cooking vegetables are proven to protect consumers from ingesting pesticide residues.

Background: Knowledge of the molecular determinants driving antigen expression is critical to design, optimize, and implement a genotyping approach on a population‐specific basis. AlthoughRHDgene variability has been extensively reported in Caucasians, Africans, and East‐Asians, it remains to be explored in Southeast Asia. Thus the molecular basis of non‐D+ blood donors was investigated in Thailand.Study Design and Methods: First, 1176 blood samples exhibiting an inconclusive or negative result by automated serological testing were collected in the 12 Regional Blood Centres of the Thai Red Cross located throughout Thailand. Second, theRHDgene was analyzed in all samples by 1) quantitative multiplex PCR of short fluorescent fragments, and 2) direct sequencing, when necessary, for identifying structural variants and single nucleotide variants, respectively.Results: Additional serological typing yielded 51 and 1125 samples with weak/partial D and D‐negative (D‐) phenotype, respectively. In the first subset, partialRHD*06.03was the most common variant allele (allele frequency: 18.6%). In the second subset, the whole deletion of the gene is largely the most frequent (allele frequency: 84.9%), followed by the AsianDELallele found in 15.6% of the samples. Eight novel alleles with various mutational mechanisms were identified.Conclusion: We report, for the first time at the national level, the molecular basis of weak/partial D and serologically D‐ phenotypes in Thai blood donors. The design and implementation of a dedicated diagnostic strategy in blood donors and patients are the very next steps for optimizing the management and supply of RBC units in Thailand.

Diabetic nephropathy, a major complication of diabetes mellitus (DM), is increasing worldwide and the large majority of patients have type 2 DM. Microalbuminuria has been used as a diagnostic marker of diabetic nephro­pathy. But owing to its insufficient sensitivity and specificity, other biomarkers are being sought. In addition, the pathophysiology of diabetic nephropathy is not fully understood and declines in renal function occur even without microalbuminuria. In this study, we investigated urinary proteins from three study groups (controls, and type 2 diabetic subjects with or without microalbuminuria). Non-targeted label-free Nano-LC QTOF analysis was conducted to discover underlying mechanisms and protein networks, and targeted label-free Nano-LC QTOF with SWATH was performed to qualify discovered protein candidates. Twenty-eight proteins were identified as candidates and functionally analyzed via String DB, gene ontology and pathway analysis. Four predictive mechanisms were analyzed: i) response to stimulus, ii) platelet activation, signaling and aggregation, iii) ECM-receptor interaction, and iv) angiogenesis. These mechanisms can provoke kidney dysfunction in type 2 diabetic patients via endothelial cell damage and glomerulus structural alteration. Based on these analyses, three proteins (kininogen-1, basement membrane-specific heparan sulfate proteoglycan core protein, and roundabout homolog 4) were proposed for further study as potential biomarkers. Our findings provide insights that may improve methods for both prevention and diagnosis of diabetic nephropathy.

The ideal therapeutic uricase (UOX) is expected to have the following properties; high expression level, high activity, high thermostability, high solubility and low immunogenicity. The latter property is believed to depend largely on sequence identity to the deduced human UOX (dH-UOX). Herein, we explored L. menadoensis uricase (LM-UOX) and found that it has 65% sequence identity to dH-UOX, 68% to the therapeutic chimeric porcine-baboon UOX (PBC) and 70% to the resurrected ancient mammal UOX. To study its biochemical properties, recombinant LM-UOX was produced in E. coli and purified to more than 95% homogeneity. The enzyme had specific activity up to 10.45 unit/mg, which was about 2-fold higher than that of the PBC. One-litre culture yielded purified protein up to 132 mg. Based on homology modelling, we successfully engineered I27C/N289C mutant, which was proven to contain inter-subunit disulphide bridges. The mutant had similar specific activity and production yield to that of wild type (WT) but its thermostability was dramatically improved. Up on storage at −20 °C and 4 °C, the mutant retained ~100% activity for at least 60 days. By keeping at 37 °C, the mutant retained ~100% activity for 15 days, which was 120-fold longer than that of the wild type. Thus, the I27C/N289C mutant has potential to be developed for treatment of hyperuricemia.

The underlying mechanism and cellular responses of bacteria against toxic cadmium ions is still not fully understood. Herein,Escherichia coliTG1 expressing hexahistidine-green fluorescent protein (His6GFP) and cells expressing polyhistidine-fused to the outer membrane protein A (His-OmpA) were applied as models to investigate roles of cytoplasmic metal complexation and metal chelation at the surface membrane, respectively, upon exposure to cadmium stress. Two-dimensional gel electrophoresis (2-DE) and two-dimensional difference in gel electrophoresis (2D-DIGE) in conjunction with mass spectrometry-based protein identification had successfully revealed the low level expression of antioxidative enzymes and stress-responsive proteins such as manganese-superoxide dismutase (MnSOD; +1.65 fold), alkyl hydroperoxide reductase subunit C (AhpC; +1.03 fold) and DNA starvation/stationary phase protection protein (Dps; −1.02 fold) in cells expressing His6GFP in the presence of 0.2 mM cadmium ions. By contrarily, cadmium exposure led to the up-regulation of MnSOD of up to +7.20 and +3.08 fold in TG1-carrying pUC19 control plasmid and TG1 expressing native GFP, respectively, for defensive purposes against Cd-induced oxidative cell damage. Our findings strongly support the idea that complex formation between cadmium ions and His6GFP could prevent reactive oxygen species (ROS) caused by interaction between Cd²⁺and electron transport chain. This coincided with the evidence that cells expressing His6GFP could maintain their growth pattern in a similar fashion as that of the control cells even in the presence of harmful cadmium. Interestingly, overexpression of either OmpA or His-OmpA inE. colicells has also been proven to confer protection against cadmium toxicity as comparable to that observed in cells expressing His6GFP. Blockage of metal uptake as a consequence of anchored polyhistidine residues on surface membrane limited certain amount of cadmium ions in which some portion could pass through and exert their toxic effects to cells as observed by the increased expression of MnSOD of up to +9.91 and +3.31 fold in case of TG1 expressing only OmpA and His-OmpA, respectively. Plausible mechanisms of cellular responses and protein mapping in the presence of cadmium ions were discussed. Taken together, we propose that the intracellular complexation of cadmium ions by metal-binding regions provides more efficiency to cope with cadmium stress than the blockage of metal uptake at the surface membrane. Such findings provide insights into the molecular mechanism and cellular adaptation against cadmium toxicity in bacteria.

Neisseria gonorrhoeae strains displaying reduced susceptibility and resistance to extended-spectrum cephalosporins (ESCs) are major public health concerns. Although resistance mechanisms of ESCs have extensively been studied, the proteome-wide investigation on the biological response to the antibiotic stress is still limited. Herein, a proteomics approach based on two-dimensional gel electrophoresis and MALDI-TOF/TOF-MS analysis was applied to investigate the global protein expression under ESC stresses of ESC-susceptible and ESC-reduced susceptible N. gonorrhoeae strains. Upon exposure to ceftriaxone, 14 and 21 proteins of ESC-susceptible and ESC-reduced susceptible strains, respectively, were shown to be differentially expressed. In the meanwhile, differential expressions of 13 and 17 proteins were detected under cefixime stress for ESC-susceptible and ESC-reduced susceptible strains, respectively. ESC antibiotics have been proven to trigger the expression of several proteins implicated in a variety of biological functions including transport system, energy metabolism, stress response and pathogenic virulence factors. Interestingly, macrophage infectivity potentiators (Ng-MIP) showed increased expression for ESC-reduced susceptible strain under ESC stress. The altered expression of Ng-MIP was found to be a unique response to ESC stresses. Our finding proposes a broad view on proteomic changes in N. gonorrhoeae in response to ESC antibiotics that provides further insights into the gonococcal antimicrobial resistance and physiological adaptation mechanism.

Among microcytic hypochromic anemias, the most common disorders are iron deficiency anemia and co-pathological conditions such as α- or β-thalassemia (α- or β-thal) traits. The aim of the present study was to determine the frequency and prevalence of iron deficiency anemia and α- or β-thal traits based on clinical laboratory data across different ethnic groups in five districts of Sindh Province, Pakistan. The present retrospective study analyzed 3 years (2012-2015) of encoded and unlinked clinical laboratory data, and identified 3030 microcytic hypochromic anemia cases. The data contained complete blood counts (CBCs) with smear morphology examinations, serum ferritin levels, and hemoglobin (Hb) electrophoreses. After reviewing the data, 994 confirmed subjects (iron deficiency anemia and α- and β-thal traits) were then selected for the present study. The prevalence of α- and β-thal traits was highest in Badin district (35.27%), while the prevalence of iron deficiency anemia was highest in Larkana district (30.73%). According to the ethnic-wise distribution, higher numbers of α- and β-thal trait cases were seen in the Sindhi ethnic group [375 (64.21%) and 283 (69.02%), respectively] than in the other ethnic groups. In addition, a higher distribution of β-thal trait cases was observed in the Sindhi ethnic group [n = 327 (56%)] in α- and β-thal cases overall. Findings from the present study strongly suggested that screening is important not only for β-thal trait but also other traits as well. However, careful monitoring of CBC parameters, including red blood cell (RBC) indices and morphology, along with clinical findings are essential to diagnose carrier cases, especially in high prevalence areas.