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The inhibition of poly(ADP-ribose) polymerases (PARPs) and ataxia telangiectasia and Rad3-related (ATR) would be an alternative approach for cancer treatments. The aim of this study is to investigate the synergy of the different combinations of PARP inhibitors (olaparib, talazoparib, or veliparib) and ATR inhibitor AZD6738. A drug combinational synergy screen that combines olaparib, talazoparib, or veliparib with AZD6738 was performed to identify the synergistic interaction, and the combination index was calculated to verify synergy. TK6 isogenic cell lines with defects in different DNA repair genes were used as a model. Cell cycle analysis, micronucleus induction, and focus formation assays of serine-139 phosphorylation of the histone variant H2AX demonstrated that AZD6738 diminished G2/M checkpoint activation induced by PARP inhibitors and allowed DNA damage-containing cells to continue dividing, leading to greater increases in micronuclei as well as double-strand DNA breaks in mitotic cells. We also found that AZD6738 was likely to potentiate cytotoxicity of PARP inhibitors in homologous recombination repair deficiency cell lines. AZD6738 sensitized more genotypes of DNA repair-deficient cell lines to talazoparib than to olaparib and veliparib, respectively. The combinational approach of PARP and ATR inhibition to enhance response to PARP inhibitors could expand the utility of PARP inhibitors to cancer patients without BRCA1/2 mutations.

Ring finger protein 43 (RNF43) is an E3 ubiquitin ligase which is well-known for its role in negative regulation of the Wnt-signaling pathway. However, the function in DNA double-strand break repairs has not been investigated. In this study, we used a lymphoblast cell line, DT40, and mouse embryonic fibroblast as cellular models to study DNA double-strand break (DSB) repairs. For this purpose, we created RNF43 knockout, RNF43-/- DT40 cell line to investigate DSB repairs. We found that deletion of RNF43 does not interfere with cell proliferation. However, after exposure to various types of DNA-damaging agents, RNF43-/- cells become more sensitive to topoisomerase II inhibitors, etoposide, and ICRF193, than wild type cells. Our results also showed that depletion of RNF43 results in apoptosis upon etoposide-mediated DNA damage. The delay in resolution of γH2AX and 53BP1 foci formation after etoposide treatment, as well as epistasis analysis with DNAPKcs, suggested that RNF43 might participate in DNA repair of etoposide-induced DSB via non-homologous end joining. Disturbed γH2AX foci formation in MEFs following pulse etoposide treatment supported the notion that RNF43 also functions DNA repair in mammalian cells. These findings propose two possible functions of RNF43, either participating in NHEJ or removing the blockage of 5' topo II adducts from DSB ends.

TRIM29 is involved in DNA double‐strand break repair but the specific roles of TRIM29 in DNA repair are not clearly understood. We proposed that TRIM29 regulates the choice of DNA DSB repair pathway by facilitating 53BP1 accumulation to promote NHEJ, possibly through RNF8 or TIP60., Tripartite motif‐containing protein 29 (TRIM29) is involved in DNA double‐strand break (DSB) repair. However, the specific roles of TRIM29 in DNA repair are not clearly understood. To investigate the involvement of TRIM29 in DNA DSB repair, we disrupted TRIM29 in DT40 cells by gene targeting with homologous recombination (HR). The roles of TRIM29 were investigated by clonogenic survival assays and immunofluorescence analyses. TRIM29 triallelic knockout (TRIM29 −/−/−/+) cells were sensitive to etoposide, but resistant to camptothecin. Foci formation assays to assess DNA repair activities showed that the dissociation of etoposide‐induced phosphorylated H2A histone family member X (ɣ‐H2AX) foci was retained in TRIM29 −/−/−/+ cells, and the formation of etoposide‐induced tumor suppressor p53‐binding protein 1 (53BP1) foci in TRIM29 −/−/−/+ cells was slower compared with wild‐type (WT) cells. Interestingly, the kinetics of camptothecin‐induced RAD51 foci formation of TRIM29 −/−/−/+ cells was higher than that of WT cells. These results indicate that TRIM29 is required for efficient recruitment of 53BP1 to facilitate the nonhomologous end‐joining (NHEJ) pathway and thereby suppress the HR pathway in response to DNA DSBs. TRIM29 regulates the choice of DNA DSB repair pathway by facilitating 53BP1 accumulation to promote NHEJ and may have potential for development into a therapeutic target to sensitize refractory cancers or as biomarker of personalized therapies.

Spinal muscular atrophy (SMA) is one of the leading causes of death in infants and young children from heritable diseases. Patients diagnosed with SMA develop symmetrical progressive muscle weakness and atrophy from degeneration of alpha motor neurons. Approximately 95% of patients have a homozygous deletion of survival motor neuron 1 (SMN1) gene in exon 7 and inherited in autosomal recessive pattern. Considering the high prevalence of SMA carrier in many population, it is possible that SMA is one of the most common autosomal recessive disorders in Thailand and Southeast Asia. In this study, we analyzed DNA from peripheral blood of 505 healthy Thai adults using quantitative PCR-based for SMN1 gene exon 7 copy number analysis. Individual samples with heterozygous deletion of SMN1 gene were confirmed with MLPA. The result identified 9 samples (1.78%) with heterozygous deletion and 39 samples as more than 2 copies of SMN1. No homozygous deletion was detected in the samples. In conclusion, we established carrier frequency of SMA in selected Thai population at 1.8% from 505 participants. The prevalence coincides with prevalence in East Asia and Caucasian population. The result could be implemented for SMA carrier screening in couples at risk in the region.

Recent developments in chemotherapy focus on target-specific mechanisms, which occur only in cancer cells and minimize the effects on normal cells. DNA damage and repair pathways are a promising target in the treatment of cancer. In order to identify novel compounds targeting DNA repair pathways, two key proteins, 53BP1 and RAD54L, were tagged with fluorescent proteins as indicators for two major double strand break (DSB) repair pathways: non-homologous end-joining (NHEJ) and homologous recombination (HR). The engineered biosensor cells exhibited the same DNA repair properties as the wild type. The biosensor cells were further used to investigate the DNA repair activities of natural biological compounds. An extract from Phyllosticta sp., the endophyte isolated from the medicinal plant Garcinia cowa Roxb. ex Choisy, was tested. The results showed that the crude extract induced DSB, as demonstrated by the increase in the DNA DSB marker &gamma, H2AX. The damaged DNA appeared to be repaired through NHEJ, as the 53BP1 focus formation in the treated fraction was higher than in the control group. In conclusion, DNA repair-based biosensors are useful for the preliminary screening of crude extracts and biological compounds for the identification of potential targeted therapeutic drugs.

Aim Vascular calcification (VC) is common among patients with chronic kidney disease (CKD) due to the strong prevalence of cardiovascular and CKD-related risk factors such as diabetes mellitus (DM), hypertension and phosphate retention. Kidney transplantation improves kidney function and abnormal mineral metabolism at the same time. It remains unclear whether kidney transplantation favourably impacts VC in the long-term. Methods The present study examined VC in 132 kidney transplant (KT) recipients who had been transplanted for longer than one year. The severity of VC was compared to 129 CKD stages 5-5D patients on a kidney transplant (KT) waiting list. Results The median KT vintage was 88 months. The prevalence of VC among KT and CKD patients were 54.5% and 62.8%, respectively, (P = 0.2). There were no differences in age, gender, body mass index (BMI), the prevalence of DM or CVD between the two groups. Among patients with calcification, a more severe degree was observed in KT recipients (P = 0.01). Aging, DM, CVD and dialysis vintage were associated with significant VC in both groups. The degree of VC in KT recipients was more pronounced than that in CKD patients among those who experienced prolonged dialysis vintage (>2 years) (P = 0.04). Among KT recipients, the severity of VC increased with the length of time after transplantation and became more substantial after 5 years. Conclusions Long-term KT recipients demonstrated a more severe degree of VC compared to matched CKD stages 5-5D patients. The severity of VC became more pronounced among those with longer transplant vintage and was in part influenced by past dialysis experience.

Background. An increased prevalence of metabolic syndrome including nonalcoholic fatty liver disease (NAFLD) was reported in psoriasis. NAFLD can progress to nonalcoholic steatohepatitis and fibrosis. Transient elastography (TE) is a noninvasive liver fibrosis assessment. We evaluated the prevalence of significant liver fibrosis or high liver stiffness measurement (LSM) using the LSM cutoff over 7 kPa and its associated factors in psoriatic patients. Methods. Subjects underwent TE and ultrasonography. Univariate and multivariate analysis were performed for the associated factors. Results. One hundred and sixty-eight patients were recruited. Three patients were excluded due to TE failure. Mean BMI was 24.8±4.7 kg/m2. NAFLD, metabolic syndrome, and diabetes were seen in 105 (63.6%), 83 (50.3%), and 31 (18.8%) patients. The total cumulative dose of methotrexate over 1.5 g was seen in 39 (23.6%) patients. Mean LSM was 5.3±2.9 kPa. High LSM was found in 18 (11.0%) patients. Waist circumference (OR: 1.24; 95% CI: 1.11–1.38; P=0.0002), diabetes (OR: 12.70; 95% CI: 1.84–87.70; P=0.010), and AST (OR: 1.08; 95% CI: 1.02–1.16; P=0.017) were associated with high LSM. Conclusion. 11% of psoriatic patients had significant liver fibrosis by high LSM. Waist circumference, diabetes, and AST level were the independent predictors.

Background Hepatitis is a common adverse effect of antituberculosis drugs. Silymarin prevented drug-induced hepatoxicity in animals with anti-oxidative mechanisms but its effect in human has been unknown. We aimed to evaluate the efficacy of silymarin for preventing antituberculosis-drug induced liver injury (antiTB-DILI) in patients with tuberculosis. Methods A double-blind randomized placebo-controlled trial was performed. Tuberculosis patients were randomly allocated to receive placebo or silymarin. The outcomes of interests were antiTB-DILI and the maximum liver enzymes at week 4. Antioxidative enzymes (i.e., superoxide dismutase (SOD), glutathione and malondialdehyde assays) were assessed. The risks of antiTB-DILI between the two groups were compared. A number need to treat was estimated. Results A total of 55 out of 70 expected numbers of patients were enrolled. There were 1/27 (3.7 %) and 9/28 (32.1 %) patients who developed antiTB-DILI in the silymarin and the placebo groups. Risk reduction was 0.28 (0.10, 0.47), i.e., receiving silymarin was 28 % at lower risk for antiTB-DILI than placebo. This led to prevention of 28 patients from being antiTB-DILI among 100 treated patients. Median (IQR) of ALT levels at week 4 in the placebo and the silymarin group were 35.0 (15, 415) IU/L and 31.5 (20, 184) IU/L (p = 0.455). The decline of SOD level at week 4 in the silymarin group was less than the placebo group (p

Background. ‘Primary’ osteoporosis has been associated with a high incidence of a renal acidification defect, incomplete renal tubular acidosis (iRTA). An acid loading test, to exclude the defect, has been recommended for inclusion in the work-up of osteoporosis. However, there is no community-based study to confirm its utility. Method. A community-based survey was conducted in the Khon Kaen province, Thailand, between January and June, 2000. We randomly enrolled 361 apparently healthy adults, 146 men and 215 women, in this study. The bone mineral densities (BMDs) of the spine and femur were determined in all subjects. The diagnosis of iRTA was based on: normal serum electrolytes and one or both of first morning urinary pH >5.5 or the failure of an acid loading test to decrease it to >5.5. Dietary diaries, serum electrolyte tests and 24 h urine collections were obtained from all iRTA subjects. Results. There were 23 (6.4%) iRTA subjects in the population studied. The age, height, weight and calcium intake were comparable between iRTA and normal subjects, as were the BMDs of spine and femur. There was no difference between the two groups in the distributions of BMD with age for either area. Multiple regression analyses of the studied population demonstrated that age, body weight, duration of menopause and gender (only for the femoral neck) were independent variables that affected BMD. Conclusion. Incomplete distal renal tubular acidosis alone was not associated with lower bone mass in this cohort. It may nevertheless be valuable to monitor serum electrolytes and BMD in patients with iRTA due to their tendency to develop intermittent metabolic acidosis.

Our previous report on bone histomorphometry in patients with distal renal tubular acidosis (dRTA) revealed decreased bone formation rate (BFR) when compared to healthy subjects. The abnormality improved significantly after alkaline therapy. The modest increase in osteoblastic surface, after correction of metabolic acidosis, could not explain the striking improvement in bone formation, suggesting additional influence of metabolic acidosis on osteoblast function and/or bone matrix mineralization. Osteoblasts and, to a lesser extent, osteoclasts synthesize and secrete bone matrix including type I collagen and various noncollagenous proteins (NCPs). Substantial evidence suggested diverse functions of NCPs related to bone formation, resorption, and mineralization. Metabolic acidosis, through its effect on bone cells, may result in an alteration in the production of NCPs. Our study examined bone histomorphometry with detailed analysis on the mineralization parameters and NCPs expression within the bone matrix of patients with dRTA before and after treatment with alkaline. Seven dRTA patients underwent bone biopsy at their initial diagnosis and again 12 months after alkaline therapy. Bone mineral density (BMD) and bone histomorphometry were obtained at baseline and after the treatment. The expression of NCPs was examined by immunohistochemistry, quantitated by digital image analysis, and reported as a percentage of area of positive staining or mineralized trabecular bone area. Alkaline therapy normalized the low serum phosphate and PTH during acidosis. The reduction in BMD at baseline improved significantly by the treatment. Bone histomorphometry demonstrated the increase in osteoid surface and volume without significant alteration after acidosis correction. In comparison to the normal subjects, osteoid thickness was slightly but insignificantly elevated. Osteoblast and osteoclast populations and their activities were suppressed. The reduction in mineral apposition rate and adjusted apposition rate were observed in conjunction with the prolongation of mineralization lag time. Alkaline therapy improved the mineralization parameters considerably. In addition to the increase in BFR relative osteoblast number after acidosis correction, osteocalcin expression in the bone matrix increased significantly from 16.7% to 22.3%. Six of seven patients had decreased osteopontin expression. In conclusion, the abnormal bone remodeling in dRTA is characterized by low turnover bone disease with some degree of defective mineralization. Alteration of NCPs expression suggested the effect of metabolic acidosis on bone cells. Alkaline therapy increased bone mass through the restoration of bone mineral balance and, perhaps, improved osteoblast function.

Potassium citrate is an alkaline agent that has been recommended for the prevention of nephrolithiasis in distal renal tubular acidosis (RTA). Information on the effectiveness and the optimal dose of potassium citrate in the correction of urinary abnormalities in pediatric distal RTA is limited, however. We conducted this study to determine the effectiveness and the optimal dose of potassium citrate for the correction of urinary abnormalities and the prevention of nephrolithiasis in children with distal RTA. Eight pediatric distal RTA patients participated in this study. The mean +/- SEM age was 9.7 +/- 1.2 years, and mean body weight was 29.1 +/- 4.7 kg. After initial evaluation, all patients were treated with increasing dosages of potassium citrate starting from 2 mEq/kg/d in three divided doses. The dosage was increased progressively in a stepwise fashion every 2 months from 2 mEq/kg/d to 3 mEq/kg/d, then to 4 mEq/kg/d. Blood and 8-hour overnight urine samples were obtained at baseline and every 2 months before increasing the dosage of potassium citrate. Urinary saturations for calcium oxalate and calcium phosphate were estimated by using Tiselius's indices. The basal urinary calcium-to-creatinine, phosphate-to-creatinine, and calcium-to-citrate ratios and urinary saturation for calcium oxalate and calcium phosphate were elevated significantly, whereas citrate-to-creatinine ratio was reduced significantly in distal RTA patients. These ratios were normalized gradually with the increasing dosage of potassium citrate. All the aforementioned abnormalities were normalized only after the dosage of potassium citrate was raised to 4 mEq/kg/d. The elevation in urinary saturation of calcium phosphate could not be normalized throughout the study, however. These results suggest that 4 mEq/kg/d of potassium citrate supplement can correct successfully most of the urinary abnormalities and the elevated urinary saturation for calcium oxalate but not for calcium phosphate in children with distal RTA. Monitoring of urinary calcium-to-creatinine ratio or citrate-to-creatinine ratio is valuable to ensure adequate potassium citrate supplementation in this group of patients.

Although calcium supplementation can cause hypercalciuria, the risk of nephrolithiasis has been shown to decrease rather than increase among subjects who had a higher calcium intake. Hypercalciuria is also a well-established side effect of calcitriol administration. However, the risk of nephrolithiasis is not well defined. The present study was undertaken to prospectively determine the effect of calcium with or without calcitriol on physicochemical risk factors associated with calcium oxalate nephrolithiasis in Thai postmenopausal women with osteoporosis. Subjects consisted of 53 Thai women more than 10 years postmenopausal who were randomly allocated to receive 750 mg of calcium carbonate supplement alone (n = 28) or 750 mg of calcium carbonate plus 0.5 microg calcitriol (n = 25) daily. Mean +/- SEM for age was 65.3+/-1.1 years, body weight 53.5+/-1.3 kg. Urine samples for biochemical assays were collected at baseline and 3 months after treatment. Supersaturation for calcium oxalate stone formation was assessed from the 24 h urine constituents by the Tiselius's index, AP(CaOx). Three months of calcium supplement alone resulted in a modest, but not significant, increase in urinary calcium (baseline, 2.90+/-0.43 mmol/day; after treatment 3.58+/-0.54 mmol/day) with no change in urinary oxalate, citrate or magnesium. In contrast, calcium together with calcitriol caused a significant increase in urinary calcium (baseline, 2.87+/-0.41 mmol/day; after treatment, 4.08+/-0.57 mmol/day; p0.05). No significant change in other urine constituents after treatment with calcium and calcitriol was detected. Therefore, AP(CaOx) did not significantly increase either after calcium alone (baseline, 1.17+/-0.39; after treatment, 1.36+/-0.28) or after calcium plus calcitriol (baseline, 1.09+/-0.17; after treatment, 1.09+/-0.19). However, after treatments, 12 subjects (23%)--6 receiving calcium supplement alone and 6 receiving calcium plus calcitriol supplement--had high AP(CaOx) values (greater than the upper limit of 95% Cl for AP(CaOx) derived from non-stone-forming Thai women). The post-treatment/baseline ratio was 3.21+/-0.74 for urinary calcium, 1.01+/-0.19 for urinary oxalate, and 2.23+/-0.42 (median 1.15) for AP(CaOx). The post-treatment/baseline ratio of calcium, but not for urinary oxalate, had a significant correlation with the post-treatment/baseline ratio of AP(CaOx). Our findings suggest that the alteration in the risk of calcium oxalate nephrolithiasis based on urinary composition is related to the alteration in urinary calcium. The risk of calcium oxalate nephrolithiasis does not increase significantly after calcium or combined calcium and calcitriol supplement in the majority of postmenopausal women with osteoporosis.

Vascular calcification (VC) is common among patients with chronic kidney disease (CKD) due to the strong prevalence of cardiovascular and CKD-related risk factors such as diabetes mellitus (DM), hypertension and phosphate retention. Kidney transplantation improves kidney function and abnormal mineral metabolism at the same time. It remains unclear whether kidney transplantation favourably impacts VC in the long-term.The present study examined VC in 132 kidney transplant (KT) recipients who had been transplanted for longer than one year. The severity of VC was compared to 129 CKD stages 5-5D patients on a kidney transplant (KT) waiting list.The median KT vintage was 88 months. The prevalence of VC among KT and CKD patients were 54.5% and 62.8%, respectively, (P = 0.2). There were no differences in age, gender, body mass index (BMI), the prevalence of DM or CVD between the two groups. Among patients with calcification, a more severe degree was observed in KT recipients (P = 0.01). Aging, DM, CVD and dialysis vintage were associated with significant VC in both groups. The degree of VC in KT recipients was more pronounced than that in CKD patients among those who experienced prolonged dialysis vintage (2 years) (P = 0.04). Among KT recipients, the severity of VC increased with the length of time after transplantation and became more substantial after 5 years.Long-term KT recipients demonstrated a more severe degree of VC compared to matched CKD stages 5-5D patients. The severity of VC became more pronounced among those with longer transplant vintage and was in part influenced by past dialysis experience.

Summary Background and objectives Renal phosphate wasting occurs early postkidney transplantation as a result of an accumulation of parathyroid hormone and fibroblast growth factor 23 from the CKD period. Serum phosphate, parathyroid hormone, and fibroblast growth factor 23 return to baseline 1 year postkidney transplantation. What happens beyond this period is unknown. Design, setting, participants, & measurements Mineral parameters were obtained from 229 kidney transplant recipients at least 1 year posttransplantation; 46 normal subjects and 202 CKD patients with similar GFR served as controls. Factors associated with phosphate metabolism were analyzed. Results Despite the reduced graft function, most kidney transplant recipients had lower serum phosphate than normal subjects accompanied by renal phosphate loss. Fibroblast growth factor 23 was mostly lower or comparable with normal subjects, whereas parathyroid hormone was elevated in most patients. Hyperparathyroidism is also more common among kidney transplant recipients compared with CKD patients. Both parathyroid hormone and fibroblast growth factor 23 showed relationships with renal phosphate excretion, but only parathyroid hormone displayed an independent association. Parathyroid hormone showed the highest area under the curve in predicting renal phosphate leak. When patients were categorized according to parathyroid hormone and fibroblast growth factor 23 levels, only subset of patients with high parathyroid hormone had an increased renal phosphate excretion. Conclusions Relatively low serum phosphate from renal phosphate leak continued to present in long-term kidney transplantation. Both parathyroid hormone and fibroblast growth factor 23 participated in renal tubular phosphate handling, but persistent hyperparathyroidism seemed to have a greater influence in this setting.

Distal renal tubular acidosis (RTA) syndrome is a condition caused by the acidification defect in collection tubule. The syndrome is characterized by a persistent hyperchloremic, normal plasma anion gap and metabolic acidosis in patients with relatively normal glomerular filtration rate (GFR). Hypokalemia is also common. Most patients are associated with recurrent calcium oxalate or calcium phosphate stone formation and nephrocalcinosis. Metabolic bone disease and growth retardation are common complications. Occasionally, patients may present with hypokalemic paralysis. The treatment aims to provide adequate alkaline therapy to neutralize the daily acid production. All the associated complications should be prevented. Monitoring of urinary parameters such as calcium-to-creatinine or citrate-to-creatinine ratio is essential to ensure appropriate dosage of alkaline therapy.

To evaluate 25 hydroxyvitamin D (25-OH-D) deficiency in a cohort ofpredialysis CKD patients and the treatment effect and safety of high dose ergocalciferol supplement in predialysis CKD.Fifty-six predialysis CKD patients who came for a regular visit at a single hospital with calculated glomerular filtration rateor =60 mL/min/1.73 m2 were screened for 25-OH-D levels. Forty-four patients with 25-OH-D deficiency were recruited into this prospective observational study that examined the effect of high dose oral ergocalciferol supplementation. After eight weeks, 37 patients completed the follow-up and biochemical parameters were reevaluated and analyzed.The mean 25-OH-D level of 56 patients was 25.6 +/- 8 ng/mL. Forty-four (78.5%) patients had 25-OH-D levels less than 30 ng/mL and four (7.1%) had severe deficiency with the level less than 15 ng/mL. High dose ergocalciferol supplement successively increased 25-OH-D levels in 35 (95%) patients. 25-OH-D levels increased significantly from 22 +/- 4.8 to 34.5 +/- 10.8 ng/mL after eight weeks (p0.001). During the study period, there were no changes in serum calcium, phosphate, and PTH. There was no other side effect associated with the treatment.25-OH-D deficiency were found in this cohort of predialysis CKD patients. Ergocalciferol was a safe and effective supplement for the 25-OH-D in predialysis CKD.

Background. Metabolic acidosis (MA) adversely affects protein and lipid metabolism as well as endocrine function. Adipose tissue communicates with the rest of the body through synthesis and release adipokines, such as leptin, adiponectin and TNF-alpha. Adiponectin enhances insulin sensitivity and possesses anti-atherogenic and anti-inflammatory properties. Circulating adiponectin correlates inversely with cardiovascular events. It is possible that MA negatively regulates adiponectin contributing to poor patient outcome. The present study investigates the effect of MA on adiponectin in vivo and in vitro. Methods. Twenty healthy female volunteers underwent a 7-day course of oral ammonium chloride (NH 4 Cl)-induced acidosis. Serum adiponectin was determined before and after NH 4 Cl ingestion. Adipocytes were differentiated from their precursors, human mesenchymal stem cells (hMSCs), in culture. Concentrated HCl was added to the media to lower pH. Adiponectin mRNA and protein were determined at 48 and 96 h by real-time RT-PCR and ELISA, respectively. Results. After a 7-day course of NH 4 Cl, serum bicarbonate decreased significantly associated with the increase in urine ammonium and titratable acid. Adiponectin decreased significantly from 10 623 to 9723 pg/mL (P < 0.005). MA suppressed adiponectin mRNA in hMSC-derived adipocytes at 48 and 96 h (P < 0.01). The amount of adiponectin released into the culture media declined corresponding to the mRNA levels (P < 0.001). MA did not affect adipocyte triglyceride or protein content. Conclusions. MA lowered circulating adiponectin through inhibition of adiponectin gene transcription in adipocytes.

To survey the urinary risk factors associated with recurrent calcium stone and the contribution of renal tubular acidosis to the prevalence of recurrent calcium stone formation in Thai recurrent stone formers.There were 86 consecutive recurrent calcium stone formers. Three-day dietary record, serum biochemical parameters, first morning urine pH, and two 24-hour urine collections were obtainedfrom each subject. Urinary risk factors for calcium stone formation were determined from the average of the 2-day urine collection. Normal controls were 34 subjects matched for aged, sex, and weight, and without a history of renal stone formation.Seven patients (8.1%) were diagnosed as incomplete renal tubular acidosis (iRTA). Among the 79 idiopathic calcium stone formers (ISF), 69.6%, 15.2%, 10.1%, 7.2% and 1.3% of patients were hypocitraturia, hypercalciuria, low urinary volume, hyperuricosuria and hyperoxaluria, respectively. The common combinations of risk factors were hypocitraturia plus low urine output (8.9%) or plus hypercalciuria (7.6%). There were significant differences between ISF and normal controls in urinary oxalate excretion (0.16 +/- 0.01 vs 0.12 +/- 0.01, p0.05), urinary calcium/citrate ratio (4.49 +/- 0.50 vs 2.83 +/- 0.34, p0.01) and ion activity product for calcium oxalate stone (0. 46 +/- 0.03 vs 0. 33 +/- 0.03, p0. 05). Urinary citrate in ISF varied directly with net alkaline absorption (r = 0.34, p0.005) and urinary potassium (r = 0.54, p0.001). There were significant correlations between urinary calcium excretion and both sodium excretion (r = 0.42, p0.001) and urea excretion (r = 0.41, p0.001) in ISE There were seven (8.1%) with incomplete renal tubular acidosis. Patients with iRTA tended to have less urinary citrate and higher calcium/citrate ratio than did ISF, but hypercalciuria was uncommon.Hypocitraturia was the most common urinary risk factor found in Thai recurrent idiopathic calcium stone formers followed by hypercalciuria and low urinary volume. Almost one-fourth of the stone formers had multiple risk factors. Hypocitraturia might result from low potassium and low alkaline intake. iRTA was common among recurrent calcium stone formers. Determination of morning urine pH should be a part of the investigations for urinary risk factors to avoid overlooking the diagnosis of iRTA.

Bone histology of distal renal tubular acidosis patients showed decreased bone formation with impaired bone matrix mineralization that is not entirely explained by an alteration in the mineral balance. Data from in vitro studies suggests a direct inhibitory effect of metabolic acidosis on osteoblast function. We investigated the effects of chronic metabolic acidosis on osteoblast differentiation from mesenchymal stem cells (MSCs). Human MSCs were allowed to differentiate into osteoblasts in culture. Concentrated hydrochloric acid was added to the medium to lower the bicarbonate concentration and pH. The expression of various osteoblastic genes and proteins and bone matrix mineralization were examined. Chronic metabolic acidosis enhanced the messenger RNA (mRNA) and protein expression of early osteoblast transcription factor, runx-2, whereas inhibiting osterix and having no effect on ATF-4. The expression of type I collagen, the most abundant bone matrix protein, was increased following the same pattern of runx-2. Likewise, metabolic acidosis slightly enhanced the expression of mature osteoblastic gene, osteocalcin. Study on mineralization revealed suppressed alkaline phosphatase mRNA and enzyme activity. Despite the augmented collagen deposit in acidic culture, bone matrix mineralization was impaired. In conclusion, chronic metabolic acidosis alters osteoblast differentiation from MSCs through its diverse effect on osteoblastic genes and proteins resulting in an impairment of bone formation.

Multiple factors associated with hypocitraturia have been identified. However, limited studies addressing the causal relationship to hypocitraturia are available. We therefore conducted this study to determine factors associated with hypocitraturia and show their causal relationship in recurrent calcium stone formers.Dietary review and 24-hour urine samples were obtained from all recurrent calcium stone formers referred for metabolic workup in the stone clinic. One month of oral potassium chloride supplementation was prescribed to stone formers to determine the causal relationship between urinary potassium and citrate levels.Eighty-three subjects, 44 men and 39 women, were recruited to participate in this study. Hypocitraturia (citrate300 mg/d [1.43 mmol/d]) was found in 50.6% of subjects. Four independent urinary variables associated with hypocitraturia were identified, including potassium level, net gastrointestinal alkaline absorption, calcium level, and titratable acid. Urinary potassium level was the strongest predictor of urinary citrate level. Hypocitraturic subjects also had lower fruit intake compared with subjects with high urinary citrate levels. Potassium chloride supplementation to a subgroup of this population (n = 58) resulted in a significant increase in urinary citrate excretion (350.73 +/- 27.25 versus 304.15 +/- 30.00 mg/d [1.67 +/- 0.13 versus 1.45 +/- 0.14 mmol/d]; P0.02), but no alteration in fractional excretion of citrate (19.7% +/- 2.7% versus 23.1% +/- 2.4%; P0.05).Hypocitraturia was found to be a common risk factor associated with recurrent calcium stone formation and low urinary potassium level, low alkaline absorption, low urinary calcium level, and high titratable acid excretion. Hypocitraturia is predominantly of dietary origin. Estimation of fruit intake should be included in the metabolic evaluation for recurrent calcium stone formation.

It has been speculated that calcium supplement in subjects with low oxalate intake might increase the risk of calcium stone formation due to an increase in calcium absorption without a significant reduction in oxalate absorption. There have been no human studies addressing specifically the effects of taking calcium supplements in populations whose dietary oxalate is low. This study was conducted to determine the effects of calcium supplements on the risk of calcium stone formation in a population with low oxalate intake. Thirty-two healthy male navy privates, 22.7 +/- 1.9 (mean +/- SD) years old, who had oxalate intake of less than 1 mmol/day, a serum creatinine of less than 150 micromol/l, and no history of renal stones, participated in the study. Dietary oxalate was controlled to be under 1 mmol/day throughout the study. Twenty-four hour urine collections for the determination of urinary constituents were obtained at baseline and after taking calcium supplements. Detection of calcium oxalate was performed to assess the risk of calcium oxalate stone formation. The urinary excretion of calcium was significantly elevated above baseline values while taking the calcium supplements (3.48 +/- 2.13 vs 5.17 +/- 2.61 mmol/d, p < 0.05) and urinary oxalate was significantly decreased when the subjects took calcium supplements compared to the corresponding baseline value (0.13 +/- 0.05 vs 0.17 +/- 0.07 mmol/d, p = 0.01). Urinary citrate was significantly elevated when the subjects took calcium supplements compared to the baseline (0.83 +/- 0.57 vs 0.64 +/- 0.39 mmol/d, p = 0.03). There was no significant alteration in the activity products of calcium oxalate while taking the calcium supplements (0.54 +/- 0.25 vs 0.57 +/- 0.22, p = 0.54). The effect of calcium supplements with meals, for the reduction of the risk of calcium stone formation, was unchanged, even in a population whose oxalate intake is rather low. Taking calcium supplements resulted in a reduction in urinary oxalates and an elevation in urinary citrates. Both alterations in urinary constituents counterbalanced the elevation in urinary calcium which resulted from the calcium supplements.

End-stage kidney disease has become an increasing burden in all regions of the world. However, limited epidemiologic data on chronic kidney disease in Southeast Asian populations are available. Therefore, a cohort study over a period of 12 yr (1985 to 1997) in 3499 employees of the Electric Generation Authority of Thailand, aged 35 to 55 yr, was conducted to determine the prevalence of decreased kidney function and risk factors associated with future development of decreased kidney function. The prevalence of decreased kidney function (GFR60 ml/min) increased from 1.7% (95% confidence interval [CI], 1.3 to 2.1) in 1985 to 6.8% (95% CI, 5.7 to 7.9) in 1997, and the prevalence of elevated serum creatinine was 6.1% (95% CI, 5.3 to 6.9) and 16.9% (95% CI, 15.3 to 18.5) in 1985 and 1997 surveys, respectively. The adjusted odds ratio for future development of decreased kidney function was 2.57 (1.0 to 6.81) for systolic hypertension (159 mmHg), 1.82 (1.12 to 2.98) for hyperuricemia (6.29 mg/dl), 1.68 (1.02 to 2.77) for elevated body mass index (24.9 kg/m(2)) compared with subjects with systolic BP140 mmHg, serum uric acid4.5 mg/dl, and body mass index 20.8 to 22.8 kg/m(2). The rising prevalence of decreased kidney function in this population resulted mainly from the increasing prevalence of the risk factors in the population. Screening to detect decreased kidney function and early intervention to modify the associated risk factors should be considered in otherwise healthy individuals. Future studies are also necessary to determine whether implementation of these measures results in a reduction of ESRD incidence in the population.

Schedule of taking calcium supplement and the risk of nephrolithiasis.BackgroundVariation in the timing of calcium supplement may affect gastrointestinal absorption of both calcium and oxalate differently and may associate with variable risk of calcium oxalate nephrolithiasis. There are few human studies addressing specifically the appropriate time for taking calcium supplement. Therefore, this study was performed to compare calcium bioavailability and the risk of calcium oxalate stone formation for calcium supplement taken with meal vs. taken at bedtime.MethodsThirty-two healthy male navy privates, 22.7 ± 1.9years old (mean ± SD), who had normal renal function (serum creatinine less than 150 umol/L) and no history of renal stone, participated in the study. The subjects were randomly allocated into two groups of 16 each. Group A took 1g of calcium carbonate with meal, 3 times/day; and group B took 3g/day of calcium carbonate at bedtime. After taking the regimens for 1week, followed by 1month of washout period, crossover between both groups was done. The diet was controlled throughout the study. Twenty-four–hour urine collections for the determination of urinary constituents were obtained at baseline and after taking both regimens of calcium supplement. Activity product for calcium oxalate was determined to assess the risk of calcium oxalate stone formation.ResultsUrinary excretions of calcium were significantly elevated above the baseline values when taking calcium supplement both with meal (3.48 ± 2.13mmol/day vs. 5.17 ± 2.61mmol/day, P < 0.05) and at bedtime (3.09 ± 1.70mmol/day vs. 5.08 ± 2.34mmol/day, P < 0.05). There was no difference between the two regimens in the urinary calcium excretions. The urinary oxalate was decreased significantly when the subjects took calcium supplement with meal compared with the corresponding baseline value (0.13 ± 0.05 vs. 0.17 ± 0.07mmol/d, P = 0.01). In contrast, there was no alteration in urinary oxalate when calcium supplement was taken at bedtime compared to the baseline values (0.15 ± 0.05mmol/day vs. 0.15 ± 0.06mmol/day, P = 0.9). Compared with the corresponding baseline values, there was no significant increase in the activity product for calcium oxalate when taking calcium with meal (0.54 ± 0.25 vs. 0.57 ± 0.22, P = 0.54), but it was increased significantly when calcium supplement was taken at bedtime (0.47 ± 0.21 vs. 0.72 ± 0.27, P < 0.01).ConclusionCalcium supplement should be taken with meal in order to avoid increasing the risk of calcium oxalate nephrolithiasis.

Cystine urinary stone is an autosomal recessive hereditary disease, frequently recurring and resisting fragmentation by Shockwave lithotripsy. As cases have never been reported before in Thailand, five cases of renal cystine stones at Ramathibodi Hospital were reported. Two were in the same family. In all cases the stones were removed by open surgery or percutaneous nephrolithotomy. Postoperatively, all the stones were analyzed by infrared spectroscopy for cystine. In two cases, cystine stones were also identified by scanning electron microscopy. Urine was analyzed for cystine by sodium cyanide-nitroprusside test, its concentration by spectrophotometry and cystine crystals were identified by the new crystal induction technique under light microscopy. By high-performance liquid chromatography (HPLC) test, urinary dibasic amino acids (ornithine, lysine, arginine) in these cases were also found to be significantly elevated. Clinical findings, diagnosis, treatment and prevention of cystine stones are reviewed.

Distal renal tubular acidosis (RTA) is a common cause of intractable calcium nephrolithiasis. In adults, the use of potassium citrate (PC) in distal RTA effectively decreases metabolic acidosis and the risk of calcium oxalate stone but it cannot decrease the risk of calcium phosphate stone. However, there is no report for the optimal dose of PC and the risk of calcium stone in distal RTA in children.To evaluate the optimal dose of PC that minimizes the risk of calcium nephrolithiasis in children with distal RTA.Prospective studyChildren who have distal RTA and were followed-up for 4 months. Patients were studied in a control phase, 1 month of PC 2 mEq/kg/day, 2 months of PC 3 mEq/kg/day and 1 month of PC 4 mEq/kg/day. The urine specimens of 41 normal children were measured for the reference value of the parameters determining the risk of calcium stone.Eight children (mean age of 10 +/- 3.7 years, female : male = 6: 2) with distal RTA were studied during the control phase and after receiving PC 2 mEq/kg/day for I month. Treatment with PC 2 mEq/kg/day was not able to normalize serum bicarbonate and caused no significant change in the urine citrate/creatinine ratio, and activity production of calcium phosphate stone but it caused a significant decrease in the urine calcium/citrate ratio. Although PC 3 mEq/kg/day for I month normalized plasma bicarbonate, only this dose given for 2 months caused a significant increase in the urine citrate/creatinine ratio and urine calcium/ citrate ratio to values that were not different from normal children, while the activity production of calcium phosphate stone did not decrease to normal level. The effect of PC 4 mEq/kg/day was similar to that of 3 mEq/kg/day.Potassium citrate 3 mEq/kg/day for 2 months effectively normalized serum bicarbonate and decreased the risk of calcium oxalate stone but this treatment was theoretically unable to reduce the risk of calcium phosphate stone in children with distal RTA.

Recent studies showed that postmenopausal women lost less bone mass when supplemented with calcium or estrogen therapy. However, the safety of the treatments in terms of the risk of calcium oxalate stone formation is unknown. We therefore conducted this study to determine the alteration in calcium oxalate supersaturation after calcium supplement or after combined calcium and estrogen therapy in postmenopausal osteoporotic women.Fifty-six postmenopausal women were enrolled in this study. All subjects were more than 10 years postmenopausal with vertebral or femoral osteoporosis by bone mineral density criteria. They were randomly allocated to receive either 625 mg of calcium carbonate (250 mg of elemental calcium) at the end of a meal three times a day (group A, n=26) or calcium carbonate in the same manner plus 0.625 mg/day of conjugated equine estrogen and 5 mg medrogestone acetate from day 1-12 each month (group B, n=30). The age (mean +/- S.E.M.) was 66.3 +/- 1.2 and 65.1 +/- 1.1 years, weight 54.1 +/- 1.2 and 55.3 +/- 2.1 kg, in group A and group B, respectively. Urine specimens (24-h) were collected at baseline and 3 months after treatment for the determination of calcium oxalate saturation by using Tiselius's index (AP(CaOx)) and calcium/citrate ratio.After 3 months of treatment, there was no significant alteration from baseline for urinary excretion of calcium, citrate and oxalate. Urinary phosphate excretion was significantly reduced (6.3 +/- 0.7 vs. 5.1 +/- 0.7 mmol/day for group A and 8.2 +/- 0.9 vs. 5.8 +/- 0.7 mmol/day for group B, P0.05), whereas net alkaline absorption was significantly elevated (10.1 +/- 3.6 vs. 20.1 +/- 4.4 meq/day for group A and 4.8 +/- 3.2 vs. 19.9 +/- 3.6 meq/day for group B, P0.05). Calcium/citrate ratio and AP(CaOx) determined at baseline were not different from the corresponding values after treatment in both groups; calcium/citrate: 10.1 +/- 3.1 vs. 10.1 +/- 2.5 for group A and 9.3 +/- 1.8 vs. 11.9 +/- 2.5 for group B and AP(CaOx): 1.1 +/- 0.1 vs. 1.3 +/- 0.2 for group A and 1.2 +/- 0.2 vs. 1.1 +/- 0.1 for group B. There were eight and nine patients with high AP(CaOx), or2, at baseline and after treatment, respectively.Calcium supplement with a meal or combined calcium supplement and estrogen therapy is not associated with a significant increased risk of calcium oxalate stone formation in the majority of postmenopausal osteoporotic patients. Determination of urinary saturation for calcium oxalate after calcium and estrogen supplements, especially at the initial phase of treatment, may be helpful in the avoidance of nephrolithiasis.

Bone mineral density and histology in distal renal tubular acidosis.BackgroundChronic metabolic acidosis in distal renal tubular acidosis (RTA) has been implicated in the pathogenesis of enhanced bone resorption and osteopenia, resulting in a loss of bone mineral content. However, histomorphometric and bone densitometric studies of patients who suffered from long-standing distal RTA have rarely been done.MethodsA cross-sectional study to determine the alterations of bone mineral density (BMD) and histology was done in 14 nonazotemic RTA patients (11 females and 3 males) who had never received alkaline therapy before enrolling into this study. The mean age was 32.7 ± 11.9 years. BMD measurements and transiliac bone biopsy were done in all patients. Blood chemistries, intact parathyroid hormone level, and a 24-hour urine collection for the determination of urinary calcium, phosphate, sodium, and potassium were obtained from the RTA patients at the time of bone biopsy. Data from 28 age-, sex-, and body mass index-matched, normal controls who were residents in the same area were also obtained.ResultsUrinary excretion of calcium was 2.05 ± 1.59 mmol/day. No patient had hypercalciuria. The serum intact parathyroid hormone level was 15.92 ± 8.48 pg/mL. RTA patients had lower BMD in most areas when compared with normal controls. There were two patients who suffered from a pathologic fracture at the femur. Bone histomorphometry from RTA patients shows a significantly decreased bone formation rate (0.02 ± 0.02 vs. 0.07 ± 0.045 μm3/μm2/day, P < 0.05), not significantly decreased osteoblastic surface (0.78 ± 1.03% vs. 2.6 ± 1.1%) and osteoclastic surface (0.05 ± 0.03 vs. 0.13 ± 0.23%), but significantly increased osteoid surface (31.47 ± 24.52 vs. 5.79 ± 4.39%, P < 0.05) and osteoid volume (2.95 ± 3.09 vs. 0.92 ± 1.05%, P < 0.05) when compared with those of normal controls. There was no difference in osteoid thickness (10.65 ± 6.10 vs. 8.69 ± 2.14 μm). Only one distal RTA patient who had a marked increase in osteoid thickness justified the diagnosis of osteomalacia.ConclusionsThis study demonstrates that low bone mass is common in distal RTA patients. Chronic metabolic acidosis results in suppression of bone formation and resorption, which in turn may contribute to the development of low bone mass in distal RTA patients. Although minor elevations in osteoid surface and osteoid volume are found among distal RTA patients, overt osteomalacia is not the predominant bone lesion.

Bone histology and bone mineral density after correction of acidosis in distal renal tubular acidosis.BackgroundThe association between chronic metabolic acidosis and alterations in bone cell functions has been demonstrated in vitro and in animal studies. However, the causal role of acidosis and the effects of alkaline therapy on bone histology and bone mineral density in chronic metabolic acidosis have never been systematically demonstrated in humans. This study was conducted to examine the alterations in bone mineral density and bone histology before and after correction of acidosis among patients with distal renal tubular acidosis (dRTA)MethodsCorrection of metabolic acidosis by potassium citrate was done in non-azotemic dRTA patients, 6 females and 4 males, who had never received long-term alkaline therapy before enrolling into this study. Blood chemistries, serum intact parathyroid hormone, and 24-hour urine collection for the determination of urinary calcium, phosphate, sodium, potassium, bone mineral density determination, and transiliac bone biopsy were done in all patients at baseline and after one year of potassium citrate therapy.ResultsSignificant elevations in serum bicarbonate (16.5 ± 3.0 vs. 24.6 ± 2.8 mEq/L, P < 0.05) and urinary potassium excretion (35.2 ± 7.9 vs. 55.4 ±3.5 mEq/L, P < 0.05) were observed after potassium citrate therapy. No significant alterations in other serum and urine electrolytes were found after the therapy. Serum intact parathyroid hormone level was also significantly elevated after one year of treatment (12.8 ± 7.3 vs. 26.2 ± 8.7 pg/mL, P < 0.05). Bone formation rate was significantly suppressed at baseline and was normalized by the treatment (0.02 ± 0.02 vs. 0.06 ± 0.03 μm3/μm2/day, P < 0.05). There were non-significant elevations in trabecular bone volume, osteoblastic and osteoclastic numbers. Bone mineral densities in dRTA patients were also significantly decreased below normal values in most studied areas at baseline and were significantly elevated at the trochanter of femur (0.677 ± 0.136 vs. 0.748 ± 0.144 g/c m2, P < 0.05) and total femur (0.898 ± 0.166 vs. 0.976 ± 0.154 g/c m2, P < 0.05) after the treatment.ConclusionsThis study demonstrates that alkaline therapy corrects abnormal bone cell function and elevates bone mineral density in dRTA patients, indicating the causal role of acidosis in the alterations of bone cell functions and reduction in bone mineral density. Parathyroid gland activity also may be involved in the adaptation of the body to chronic metabolic acidosis.