Analysis of SIRT1 Gene SNPs and Clinical Characteristics in Medication-Related Osteonecrosis of the Jaw.

Certain genetic factors, including single-nucleotide polymorphisms (SNPs) in the SIRT1 gene, have been linked to medication-related osteonecrosis of the jaw (MRONJ). This study examined four SNPs in the SIRT1 gene and implemented multivariate statistical analysis to analyze genetic and clinical factors in MRONJ patients. Genomic DNA was isolated from peripheral blood samples of 63 patients of European origin treated for MRONJ, and four SNP genotypes in the gene encoding the SIRT-1 protein were determined by Sanger sequencing. The allele frequencies measured in the MRONJ population were compared with allele frequencies measured in the European population in the National Center for Biotechnology Information Allele Frequency Aggregator (NCBI ALFA) database. Genetic and clinical factors were examined with multivariate statistical analysis. A C:A allele distribution ratio of 77.8:22.2 was measured in the rs932658 SNP. In the ALFA project, a C:A allele distribution ratio of 59.9:40.1 was detected in the European population, which was found to be a significant difference (p = 4.5 × 10-5). Multivariate statistical analysis revealed a positive correlation (0.275) between the genotype of SNP rs932658 and the number of stages improved during appropriate MRONJ therapy. It is concluded that allele A in SNP rs932658 in the SIRT1 gene acts as a protective factor in MRONJ.

Genome-wide Association Study Identified Chromosome 8 Locus Associated with Medication-Related Osteonecrosis of the Jaw.

Medication-related osteonecrosis of the jaw (MRONJ) is a rare but serious drug-related adverse event. To identify pharmacogenomic markers of MRONJ associated with bisphosphonate therapy, we conducted a genomewide association study (GWAS) meta-analysis followed by functional analysis of 5,008 individuals of European ancestry treated with bisphosphonates, which includes the largest number of MRONJ cases to date (444 cases and 4,564 controls). Discovery GWAS was performed in randomly selected 70% of the patients with cancer and replication GWAS was performed in the remaining 30% of the patients with cancer treated with intravenous bisphosphonates followed by meta-analysis of all 3,639 patients with cancer. GWAS was also performed in 1,369 patients with osteoporosis treated with oral bisphosphonates. The lead single-nucleotide polymorphism (SNP), rs2736308 on chromosome 8, was associated with an increased risk of MRONJ with an odds ratio (OR) of 2.71 and 95% confidence interval (CI) of 1.90-3.86 (P = 3.57*10-8 ) in the meta-analysis of patients with cancer. This SNP was validated in the MRONJ GWAS in patients with osteoporosis (OR: 2.82, 95% CI: 1.55-4.09, P = 6.84*10-4 ). The meta-analysis combining patients with cancer and patients with osteoporosis yielded the same lead SNP rs2736308 on chromosome 8 as the top SNP (OR: 2.74, 95% CI: 2.09-3.39, P = 9.65*10-11 ). This locus is associated with regulation of the BLK, CTSB, and FDFT1 genes, which had been associated with bone mineral density. FDFT1 encodes a membrane-associated enzyme, which is implicated in the bisphosphonate pathway. This study provides insights into the potential mechanism of MRONJ.

SIRT1 Gene SNP rs932658 Is Associated With Medication-Related Osteonecrosis of the Jaw.

Medication-related osteonecrosis of the jaw (MRONJ) is a rare but serious adverse drug reaction. Our previous whole-exome sequencing study found SIRT1 intronic region single-nucleotide polymorphism (SNP) rs7896005 to be associated with MRONJ in cancer patients treated with intravenous (iv) bisphosphonates (BPs). This study aimed to identify causal variants for this association. In silico analyses identified three SNPs (rs3758391, rs932658, and rs2394443) in the SIRT1 promoter region that are in high linkage disequilibrium (r2 > 0.8) with rs7896005. To validate the association between these SNPs and MRONJ, we genotyped these three SNPs on the germline DNA from 104 cancer patients of European ancestry treated with iv BPs (46 cases and 58 controls). Multivariable logistic regression analysis showed the minor alleles of these three SNPs were associated with lower odds for MRONJ. The odds ratios (95% confidence interval) and p values were 0.351 (0.164-0.751; p = 0.007) for rs3758391, 0.351 (0.164-0.751; p = 0.007) for rs932658, and 0.331 (0.157-0.697; p = 0.0036) for rs2394443, respectively. In the reporter gene assays, constructs containing rs932658 with variant allele A had higher luciferase activity than the reference allele, whereas constructs containing SNP rs3758391 and/or rs2394443 did not significantly affect activity. These results indicate that the promoter SNP rs932658 regulates the expression of SIRT1 and presumably lowers the risk of MRONJ by increasing SIRT1 expression. © 2020 American Society for Bone and Mineral Research (ASBMR).

Genetic variants of VDR and CYP2R1 affect BMI independently of serum vitamin D concentrations.

BACKGROUND: Vitamin D metabolism and obesity have been linked by several studies, however the reason for this association is unclear. Our objective was to investigate potential correlations between genetic variants in key enzymes of vitamin D metabolism and the body mass index on a representative and random sample of Hungarian adults. METHODS: Altogether 462 severely vitamin D deficient individuals were studied at the end of winter in order to decrease environmental and maximize any relevant genetic effect. Furthermore, participants with lifestyle factors known to affect vitamin D homeostasis were also excluded. We selected 23 target SNPs in five genes that encode key proteins of vitamin D metabolism (NADSYN1, GC, CYP24A1, CYP2R1, VDR). RESULTS: Variants in 2 genetic polymorphisms; rs2853564 (VDR) and rs11023374 (CYP2R1) showed a significant association with participants' BMI. These associations survived further adjustment for total-, free-, or bioactive-25(OH) vitamin D levels, although the variance explained by these 2 SNPS in BMI heterogeneity was only 3.2%. CONCLUSION: Our results show two novel examples of the relationship between genetics of vitamin D and BMI, highlighting the potential role of vitamin D hormone in the physiology of obesity.

Prenatal Diagnosis of 4q Terminal Deletion and Review of the Literature.

Terminal deletion of chromosome 4 (4q deletion syndrome) is a rare genetic condition that is characterized by a broad clinical spectrum and phenotypic variability. Diagnosis of the distinct condition can be identified by conventional chromosome analysis and small deletions by novel molecular cytogenetic methods such as microarray comparative genome hybridization (aCGH). Prenatal diagnosis is challenging; to date 10 cases have been described. We report a prenatally diagnosed case of de novo 4q deletion syndrome confirmed by conventional karyotyping and FISH due to an elevated combined risk for Down syndrome and prenatal ultrasound findings. aCGH validated the diagnosis and offered exact characterization of the disorder. Cytogenetic and microarray results described a 4q32.1qter terminal deletion of the fetus. Prenatal ultrasound detected multiple nonstructural findings (micrognathia, choroid plexus cysts, echogenic fetal bowel, short femur, and cardiac axis deviation). Pregnancy was terminated at 20 weeks. In addition to the index patient, we reviewed the 10 prenatally published cases of 4q deletion syndrome in the literature and compared these with our results. We summarize the patients' characteristics and prenatal clinical findings. Alterations of maternal serum biochemical factors, an elevated combined risk for trisomies, and distinct ultrasonographic findings can often be observed in cases of prenatal 4q deletion syndrome and may facilitate the otherwise difficult prenatal diagnosis.

Long-term selective estrogen receptor-beta agonist treatment modulates gene expression in bone and bone marrow of ovariectomized rats.

Gonadal hormones including 17ß-estradiol exert important protective functions in skeletal homeostasis. However, numerous details of ovarian hormone deficiency in the common bone marrow microenvironment have not yet been revealed and little information is available on the tissue-specific acts either, especially those via estrogen receptor beta (ERß). The aim of the present study was therefore to examine the bone-related gene expression changes after ovariectomy (OVX) and long-term ERß agonist diarylpropionitrile (DPN) administration. We found that OVX produced strong and widespread changes of gene expression in both femoral bone and bone marrow. In the bone out of 22 genes, 20 genes were up- and 2 were downregulated after OVX. It is noteworthy that DPN restored mRNA expression of 10 OVX-induced changes (Aldh2, Col1a1, Daam1, Fgf12, Igf1, Il6r, Nfkb1, Notch1, Notch2 and Psen1) suggesting a modulatory role of ERß in bone physiology. In bone marrow, out of 37 categorized genes, transcription of 25 genes were up- and 12 were downregulated indicating that the marrow is highly responsive to gonadal hormones. DPN modestly affected transcription, only expression of two genes (Nfatc1 and Tgfb1) was restored by DPN action. The PI3K/Akt signaling pathway was the most affected gene cluster following the interventions in bone and bone marrow, as demonstrated by canonical variates analysis (CVA). We suggested that our results contribute to a deeper understanding of alterations in gene expression of bone and bone marrow niche elicited by ERß and selective ERß analogs.

Impact of genetic influence on serum total- and free 25-hydroxyvitamin-D in humans.

Serum 25-hydroxyvitamin D /25OHD/ levels in humans are determined primarily by environmental factors such as UV-B radiation and diet, including vitamin D intake. Although some genetic determinants of 25OHD levels have been shown, the magnitude of this association has not yet been clarified. The present study evaluates the genetic contribution to total- /t-25OHD/ and free-25OHD /f-25OHD/ in a representative sample of the Hungarian population (n = 462). The study was performed at the end of winter to minimize the effect of sunlight, which is a major determinant of serum vitamin D levels. Single nucleotide polymorphisms (SNPs) of five genes playing major roles in vitamin D metabolism were investigated (NADSYN1, DHCR7, GC, CYP2R1 and CYP24A1). The selected SNPs account for 13.1% of the variance of t-25OHD levels. More than half of the genetic effect on t-25OHD levels was explained by two polymorphisms (rs7935125 in NADSYN1 and rs2762941 in CYP24A1), which had not previously been investigated with respect to vitamin D metabolism. No SNPs exhibited association with f-25OHD levels. Unexpectedly, SNPs that showed univariate associations with vitamin D binding protein (DBP) levels were not associated with f-25OHD levels questioning the biological significance of these polymorphisms. The present study shows that t-25OHD levels are significantly influenced by genetic factors, however, the clinical significance of this observation remains to be defined, as variation in f-25OHD levels are marginally explained by genetic effects.

Next-generation sequencing of common osteogenesis imperfecta-related genes in clinical practice.

Next generation sequencing (NGS) is a rapidly developing area in genetics. Utilizing this technology in the management of disorders with complex genetic background and not recurrent mutation hot spots can be extremely useful. In this study, we applied NGS, namely semiconductor sequencing to determine the most significant osteogenesis imperfecta-related genetic variants in the clinical practice. We selected genes coding collagen type I alpha-1 and-2 (COL1A1, COL1A2) which are responsible for more than 90% of all cases. CRTAP and LEPRE1/P3H1 genes involved in the background of the recessive forms with relatively high frequency (type VII and VIII) represent less than 10% of the disease. In our six patients (1-41 years), we identified 23 different variants. We found a total of 14 single nucleotide variants (SNV) in COL1A1 and COL1A2, 5 in CRTAP and 4 in LEPRE1. Two novel and two already well-established pathogenic SNVs have been identified. Among the newly recognized mutations, one results in an amino acid change and one of them is a stop codon. We have shown that a new full-scale cost-effective NGS method can be developed and utilized to supplement diagnostic process of osteogenesis imperfecta with molecular genetic data in clinical practice.

PREDICTIVE VALUE OF SOMATIC MUTATIONS FOR THE DEVELOPMENT OF MALIGNANCY IN THYROID NODULES BY CYTOPATHOLOGY.

OBJECTIVE: The purpose of our prospective longitudinal study was to evaluate the predictive efficacy of genetic testing for malignancies in fine-needle aspiration biopsy samples that are cytologically benign at the time of biopsy. METHODS: A total of 779 aspirated cytological samples collected from thyroid nodules of 626 patients were included in a 3-year follow-up study. Consecutive patients with cytologically benign thyroid nodules by the Bethesda System for Reporting Thyroid Cytopathology were enrolled in the study. At enrollment, somatic 1-point nucleotide polymorphisms of BRAF and RAS family genes were tested by melting-point analysis, while RET/PTC and PAX8/PPAR-gamma rearrangements were examined by real-time polymerase chain reaction. The genetic test was considered to be positive if a somatic mutation was found. Malignant cytopathologic diagnoses were confirmed by histopathology. RESULTS: In samples collected from 779 thyroid nodules, there were 39 BRAF, 33 RAS mutations, and 1 RET/PTC rearrangements found at the beginning of the study. No PAX8/PPAR-gamma rearrangement was identified. There were 52 malignant thyroid tumors removed during follow-up, out of which 24 contained a somatic mutation. The specificity of the presence of somatic mutations for malignancies was as high as 93.3%, and sensitivity was 46.2%. The negative predictive value of genetic testing reached 96.0%. CONCLUSION: Our results show that our set of genetic tests can predict the appearance of malignancy in benign thyroid nodules (at the beginning of follow-up) with high specificity and strong negative predictive value. ABBREVIATIONS: BRAF = v-raf murine sarcoma viral oncogene homolog B1 FLUS = follicular lesion of undetermined significance FNAB = fine-needle aspiration biopsy FTC = follicular thyroid carcinoma HRAS = homologous to the oncogene from the Harvey rat sarcoma virus KRAS = homologous to the oncogene from the Kirsten rat sarcoma virus NRAS = first isolated from a human neuroblastoma/neuroblastoma RAS = viral oncogene homolog PAX8 = paired box 8 PCR = polymerase chain reaction PPAR-gamma = peroxisome proliferator-activated receptor gamma PTC = papillary thyroid carcinoma RAS = rat sarcoma RET = rearranged during transfection tyrosine-kinase proto-oncogene SM = somatic mutation SNP = single-nucleotide polymorphism.

Genetic Alterations in Hungarian Patients with Papillary Thyroid Cancer.

The incidence of thyroid cancers is increasing worldwide. Some somatic oncogene mutations (BRAF, NRAS, HRAS, KRAS) as well as gene translocations (RET/PTC, PAX8/PPAR-gamma) have been associated with the development of thyroid cancer. In our study, we analyzed these genetic alterations in 394 thyroid tissue samples (197 papillary carcinomas and 197 healthy). The somatic mutations and translocations were detected by Light Cycler melting method and Real-Time Polymerase Chain Reaction techniques, respectively. In tumorous samples, 86 BRAF (44.2%), 5 NRAS (3.1%), 2 HRAS (1.0%) and 1 KRAS (0.5%) mutations were found, as well as 9 RET/PTC1 (4.6%) and 1 RET/PTC3 (0.5%) translocations. No genetic alteration was seen in the non tumorous control thyroid tissues. No correlation was detected between the genetic variants and the pathological subtypes of papillary cancer as well as the severity of the disease. Our results are only partly concordant with the data found in the literature.

Strong effect of SNP rs4988300 of the LRP5 gene on bone phenotype of Caucasian postmenopausal women.

The purpose of this study was to identify relationships between single nucleotide polymorphisms (SNPs) in the genes of the Wnt pathway and bone mineral density (BMD) of postmenopausal women. We chose this pathway due to its importance in bone metabolism that was underlined in several studies. DNA samples of 932 Hungarian postmenopausal women were studied. First, their BMD values at different sites (spine, total hip) were measured, using a Lunar Prodigy DXA scanner. Thereafter, T-score values and the patients' body mass indices (BMIs) were calculated, while information about the fracture history of the sample population was also collected. We genotyped nine SNPs of the following three genes: LRP5, GPR177, and SP7, using a Sequenom MassARRAY Analyzer 4 instrument. The genomic DNA samples used for genotyping were extracted from the buccal mucosa of the subjects. Statistical analyses were carried out using the SPSS 21 and R package. The results of this analysis showed a significant association between SNP rs4988300 of the LRP5 gene and total hip BMD values. We could not reveal any associations between the markers of GPR177, SP7, and bone phenotypes. We found no effect of these genotypes on fracture risk. We could demonstrate a significant gene-gene interaction between two SNPs of LRP5 (rs4988300 and rs634008, p = 0.009) which was lost after Bonferroni correction. We could firmly demonstrate a significant association between rs4988300 of the LRP5 gene and bone density of the hip on the largest homogeneous postmenopausal study group analyzed to date. Our finding corroborates the relationship between LRP5 genotype and bone phenotype in postmenopausal women, however, the complete mechanism of this relationship requires further investigations.

Polymorphisms of CSF1 and TM7SF4 genes in a case of mild juvenile Paget's disease found using next-generation sequencing.

Juvenile Paget's disease (JPD) is a rare autosomal-recessive condition. It is diagnosed in young children and characterized by a generalized increase in bone turnover, bone pain, and skeletal deformity. Our patient was diagnosed after a pathological fracture when she was 11 years old. When we first examined her at the age of 30 she had bone pain and deformity in both the femur and tibia. Serum alkaline phosphatase (ALP) level, radiology, bone scintigraphy, and densitometry were monitored. Next generation sequencing (NGS) technology, namely semiconductor sequencing, was used to determine the genetic background of JPD. Seven target genes and regions were selected and analyzed after literature review (TM7SF4, SQSTM1, TNFRSF11A, TNFRSF11B, OPTN, CSF1, VCP). No clear pathogenic mutation was found, but we detected missense polymorphisms in CSF1 and TM7SF4 genes. After treatment with zoledronic acid, infusion bone pain and ALP level decreased. We can conclude that intravenous zoledronic acid therapy is effective and safe for suppressing bone turnover and improving symptoms in JPD, but the long-term effects on clinical outcomes are unclear. Our findings also suggest that NGS may help explore the pathogenesis and aid the diagnosis of JPD.

Rapid and cost effective screening of breast and ovarian cancer genes using novel sequence capture method in clinical samples.

BRCA1 and BRCA2 are two well-known genes in the background of hereditary breast and ovarian cancer. There is also evidence that several other genes play an important role in the pathogenesis of these two malignancies. Latest population-scaled studies showed that certain mutations in different genes could cause similar risk elevation like BRCA2 mutations. In this study we present a new method to analyse the risk assessment of women to breast and ovarian cancer. Using Haloplex, a novel sequence capture method combined with next-generation sequencing we were able to perform rapid and cost-effective screening of 16 genes that could be associated with an increased risk of breast and ovarian cancer. The rapid and cost effective analysis of this 16-gene cohort can reveal the genetic background of approximately 30 % of hereditary and familiar cases of breast and ovarian cancers. Thus, it opens up a new and high-throughput approach with fast turnaround time to the genetic diagnostics of these disorders and may be helpful to investigate other familial genetic disorders as well.

Fast and robust next-generation sequencing technique using ion torrent personal genome machine for the screening of neurofibromatosis type 1 (NF1) gene.

Neurofibromatosis type 1 (NF1) gene exhibits one of the highest spontaneous mutation rates in the human genome. Identification of the NF1 mutation is challenging because the NF1 gene is very large and complex, lacking mutational "hot spots." There is no clustering of mutations, there are several pseudogenes, and a wide spectrum of different types of mutation has been recognized. To date, NF1 mutations or deleted regions have been detected with a number of techniques. With the appearance of next-generation sequencing (NGS) machines, molecular biology is in a new revolutionary phase. Our aim was to work out a method to use the high-throughput NGS machine, Ion Torrent PGM, in diagnostic settings for neurofibromatosis type 1. In our examination, we could reveal 21 distinct variations in NF1 gene in seven patients. This is an absolutely new method for exploring the genetic background of neurofibromatosis type 1 exhibiting the extremely high throughput of NGS in a diagnostic setting.

CYP24A1 inhibition facilitates the anti-tumor effect of vitamin D3 on colorectal cancer cells.

AIM: The effects of vitamin D3 have been investigated on various tumors, including colorectal cancer (CRC). 25-hydroxyvitamin-D3-24-hydroxylase (CYP24A1), the enzyme that inactivates the active vitamin D3 metabolite 1,25-dihydroxyvitamin D3 (1,25-D3), is considered to be the main enzyme determining the biological half-life of 1,25-D3. During colorectal carcinogenesis, the expression and concentration of CYP24A1 increases significantly, suggesting that this phenomenon could be responsible for the proposed efficacy of 1,25-D3 in the treatment of CRC. The aim of this study was to investigate the anti-tumor effects of vitamin D3 on the human CRC cell line Caco-2 after inhibition of the cytochrome P450 component of CYP24A1 activity. METHODS: We examined the expression of CYP24A1 mRNA and the effects of 1,25-D3 on the cell line Caco-2 after inhibition of CYP24A1. Cell viability and proliferation were determined by means of sulforhodamine-B staining and bromodeoxyuridine incorporation, respectively, while cytotoxicity was estimated via the lactate dehydrogenase content of the cell culture supernatant. CYP24A1 expression was measured by real-time reverse transcription polymerase chain reaction. A number of tetralone compounds were synthesized to investigate their CP24A1 inhibitory activity. RESULTS: In response to 1,25-D3, CYP24A1 mRNA expression was enhanced significantly, in a time- and dose-dependent manner. Caco-2 cell viability and proliferation were not influenced by the administration of 1,25-D3 alone, but were markedly reduced by co-administration of 1,25-D3 and KD-35, a CYP24A1-inhibiting tetralone. Our data suggest that the mechanism of action of co-administered KD-35 and 1,25-D3 does not involve a direct cytotoxic effect, but rather the inhibition of cell proliferation. CONCLUSION: These findings demonstrate that the selective inhibition of CYP24A1 by compounds such as KD-35 may be a new approach for enhancement of the anti-tumor effect of 1,25-D3 on CRC.

Gene expression changes in femoral head necrosis of human bone tissue.

Osteonecrosis of the femoral head (ONFH) is the result of an interruption of the local circulation and the injury of vascular supply of bone. Multiple factors have been implicated in the development of the disease. However the mechanism of ischemia and necrosis in non-traumatic ONFH is not clear. The aim of our investigation was to identify genes that are differently expressed in ONFH vs. non-ONFH human bone and to describe the relationships between these genes using multivariate data analysis. Six bone tissue samples from ONFH male patients and 8 bone tissue samples from non-ONFH men were examined. The expression differences of selected 117 genes were analyzed by TaqMan probe-based quantitative real-time RT-PCR system. The significance test indicated marked differences in the expression of nine genes between ONFH and non-ONFH individuals. These altered genes code for collagen molecules, an extracellular matrix digesting metalloproteinase, a transcription factor, an adhesion molecule, and a growth factor. Canonical variates analysis demonstrated that ONFH and non-ONFH bone tissues can be distinguished by the multiple expression profile analysis of numerous genes controlled via canonical TGFB pathway as well as genes coding for extracellular matrix composing collagen type molecules. The markedly altered gene expression profile observed in the ONFH of human bone tissue may provide further insight into the pathogenetic process of osteonecrotic degeneration of bone.

The role of vitamin D, estrogen, calcium sensing receptor genotypes and serum calcium in the pathogenesis of prostate cancer.

INTRODUCTION: Prostate cancer is the second leading cause of cancer death among men in developed countries. Estrogen receptor-alpha (ER-α), vitamin D receptor (VDR), and the calcium-sensing receptor (CaSR), partly through their effects on calcium levels are implicated in the proliferation and carcinogenesis in the prostate gland. VDR, ER-α and CaSR genes show polymorphisms in humans that appear to have clinical significance in many pathological conditions, such as prostate cancer. Our aim was to evaluate the role of ER-α (PvuII, XbaI), VDR (BsmI) and CaSR (A986S) gene polymorphisms and serum calcium levels in the pathogenesis of prostate cancer. MATERIAL AND METHODS: Two hundred four patients with prostate cancer and 102 healthy controls were recruited into a hospital-based case control study. After genotyping, the relationship between the individual genotypes and prostate cancer was investigated. RESULTS: Both the ER-α XbaI and the VDR BsmI polymorphisms were significantly related to the risk of prostate cancer. An age adjusted logistic regression limited to controls and patients not receiving bisphosphonate therapy showed that higher corrected serum calcium and the VDR Bb/BB genotypes independently increased the risk of prostate cancer. CONCLUSIONS: ER-α XbaI and VDR BsmI genetic polymorphisms had a significant association with the risk of prostate cancer. Both VDR BsmI genotypes and serum calcium levels were independently related to the risk of prostate cancer, suggesting an influence of VDR on the development of this malignancy.

The protective role of bone morphogenetic protein-8 in the glucocorticoid-induced apoptosis on bone cells.

One of the side effects associated with glucocorticoid therapy is glucocorticoid-induced bone loss. Glucocorticoids partly detain bone formation via the inhibition of osteoblastic function, however, the exact mechanism of this inhibition remains elusive. In this study, we examined the effect of dexamethasone, an active glucocorticoid analogue, on cell viability and expression of bone remodelling-related genes in primary mouse calvarial and cloned MC3T3-E1 osteoblasts. Using sensitive biochemical assays, we demonstrated the apoptotic effect of dexamethasone on osteoblastic cells. Then, utilizing Taqman probe-based quantitative RT-PCR technology, gene expression profiles of 111 bone metabolism-related genes were determined. As a result of dexamethasone treatment we have detected significant apoptotic cell death, and six genes, including Smad3, type-2 collagen α-1, type-9 collagen α-1, matrix metalloproteinase-2, bone morphogenetic protein-4 and bone morphogenetic protein-8 showed (BMP-8) significant changes in their expression on a time- and concentration-dependent manner. BMP-8, (a novel player in bone-metabolism) exhibited a two orders of magnitude elevation in its mRNA level and highly elevated protein concentration by Western blot in response to dexamethasone treatment. The knockdown of BMP-8 by RNA interference significantly increased dexamethasone-induced cell death, confirming a protective role for BMP-8 in the glucocorticoid-induced apoptosis of osteoblasts. Our results suggest that BMP-8 might be an essential player in bone metabolism, especially in response to glucocorticoids.

Association between the malnutrition-inflammation score and post-transplant anaemia.

BACKGROUND: Post-transplant anaemia (PTA) is common and is associated with adverse consequences. The protein-energy wasting (PEW) syndrome is associated with erythropoietin resistance in patients on maintenance dialysis. We assessed the association between PEW and PTA in a large prevalent cohort of stable kidney-transplanted patients. METHODS: Data from 942 prevalent kidney-transplanted patients were analysed. Socio-demographic parameters, laboratory results, transplantation-related data and medication were obtained from the charts. Biomarkers reflecting nutritional status and inflammation [serum leptin, albumin, interleukin-6 (IL-6), tumour necrosis factor-α (TNF-α) and C-reactive protein] were measured. Anthropometric measures and the malnutrition-inflammation score (MIS) were also tabulated. Anaemia was defined according to the guidelines of the American Society of Transplantation. RESULTS: Mean age was 51 ± 13 years, 57% were males and 22% had diabetes. The prevalence of PTA was 33%. The haemoglobin (Hb) level significantly and negatively correlated with the MIS (rho = - 0.316), marginally with serum TNF-α (rho = - 0.079) and serum IL-6 (rho = - 0.075) and positively with serum transferrin (r = 0.298), serum albumin (r = 0.274), abdominal circumference (r = 0.254) and serum leptin (rho = - 0.152), P < 0.05 for all. In a multivariable linear regression model, MIS was independently associated with Hb (beta = - 0.118, P = 0.004) in patients with estimated glomerular filtration rate (eGFR) lower than or equal to 60 mL/min/1.73 m(2), but not in patients with higher eGFR. CONCLUSIONS: The MIS is independently associated with PTA in the kidney-transplanted population with eGFR lower than or equal to 60 mL/min/1.73 m(2).