Collagen Type 1 Alpha 1 Research Articles

Imatinib mesylate for children with dermatofibrosarcoma protuberans (DFSP)

Dermatofibrosarcoma protuberans (DFSP) is a rare malignant soft tissue tumor in children. DFSP is characterized by a specific fusion of the platelet-derived growth factor beta (PDGFbeta) with the collagen type 1alpha1 (COL1alpha1) gene which renders these tumors responsive to targeted therapy with tyrosine kinase inhibitors, such as imatinib mesylate, as is reported in adults. In the current report, we describe the first small pediatric DFSP series, in which response to imatinib mesylate contributed to successful treatment outcome.

Preliminary Evidence of an Association Between COL1A2 Polymorphisms and Dental Fluorosis in a Population with High Fluoride Exposure

COL1A2 gene polymorphisms (Pvu II and Rsa I), serum calciotropic hormone levels, and dental fluorosis. Huang H, Ba Y, Cui L, Cheng X, Zhu J, Zhang Y, et al. Community Dent Oral Epidemiol 2008;36(6):517-22. Deborah V. Dawson, PhD, ScM PURPOSE/QUESTION: To assess the relationships among polymorphisms in the collagen type 1 alpha 2 (COL1A2) gene, serum calciotropic hormone levels, and dental fluorosis. Government-Department of Science and Technology of Henan Province, China Case-control study Level 2: Limited-quality, patient-oriented evidence Not applicable.

PDGFBquantification is a useful tool in the diagnosis of dermatofibrosarcoma protuberans: a study of 10 cases

Genetic aberrations involving the platelet-derived growth factor-beta (PDGFB) gene and the collagen type 1 alpha1 (COL1A1) gene have been implicated in the pathogenesis of dermatofibrosarcoma protuberans (DFSP), a slow-growing and locally infiltrative dermal tumour. To investigate the genetic rearrangements in 10 patients who presented with a clinical diagnosis of DFSP. Total RNA was obtained from frozen sections and sections embedded in paraffin wax, and used for direct sequencing of the cDNA produced from reverse transcription (RT) PCR. The expression of PDFGB mRNA in each of these cases was also examined. Of the 10 samples examined, 9 had the COL1A1/PDGFB fusion transcript by DNA sequencing. The sequenced products showed that there was a fusion between the end of exons 6, 8, 29 (two cases), 38, 25 or 47 (three cases) and the start of exon 2 of the PDGFB gene. Quantitative RT-PCR identified all samples as having significantly higher expression of the PDGFB gene compared with normal skin or dermatofibroma. Detection of the COL1A1/PDGFB fusion transcript may be important for the diagnosis of DFSP. Furthermore, relative PDGFB gene quantification by real-time PCR may also provide a good diagnostic tool when other methods fail to give conclusive results.

CPG HYPERMETHYLATION OF COL1A2 CONTRIBUTES TO PROLIFERATION AND MIGRATION ACTIVITY OF BLADDER CANCER

INTRODUCTION AND OBJECTIVE: Gene silencing due to aberrant DNA methylation plays an important role in carcinogenesis. For genome-wide screening for methylated genes in bladder cancer (BC), we performed cDNA microarray analysis of the BC cell line (BOY) treated with demethylating agent (5-Aza-dC). Collagen type 1 alpha 2 (COL1A2) gene was identified as the most up-regulated one of the 30,144 genes screened. We hypothesize that inactivation of the COL1A2 gene through CpG methylation contributes to proliferation and migration activity of human BC by changing extra-cellular matrix (ECM). METHODS: To test this hypothesis, we extracted DNA and RNA from 67 BC samples and 10 normal bladder epitheliums (NBEs) from cystectomy and transurethral resection. The CpG hypermethylation of COL1A2 promoter was analyzed by quantitative methylation-specific PCR (QMSP) and confirmed by bisulfite-modified DNA sequencing. Real-time RT-PCR was carried out to evaluate COL1A2 mRNA expression. We established a stable COL1A2 transfectant for gain of function studies, for which cellular proliferation and migration were examined by MTT assay and wound healing assay, respectively. The protein secretion of type 1 collagen in culture medium was detected by immunoblot. RESULTS: Bisulfite DNA sequencing demonstrated that the promoter hypermethylation of COL1A2 was a frequent event in clinical BCs. The methylation index was significantly higher in the 67 BCs than in the10 NBEs (42.3 ± 28.9 vs. 0.07 ± 0.01, p = 0.0011). Conversely, COL1A2 mRNA expression was significantly lower in the BCs than in the NBEs (15.8 ± 3.8 vs. 32.0 ± 22.0, p = 0.0052). In vitro, after 5-aza-dC treatment, the COL1A2 mRNA expression was markedly increased in BOY. Our cell proliferation assays consistently demonstrated a significant growth inhibition in the COL1A2 transfectant compared with control and wild-type BOY cells (on day 5; 0.791 ± 0.020, 1.056 ± 0.034, and 1.107 ± 0.027, respectively, p < 0.0001). Wound healing assays also showed a significant wound healing inhibition in the COL1A2 transfectant compared to the counterparts (% of wound closure; 75.5 ± 0.2, 94.3 ± 2.9, and 96.9 ± 3.1, respectively, p = 0.0016). Immunoblot demonstrated that type 1 collagen protein markedly increased in culture medium of the COL1A2 transfectant. CONCLUSIONS: Our data suggest that inactivation of the COL1A2 gene through CpG methylation may change an ECM binding on epithelial collagen and contributes to proliferation and migration activity of human bladder cancer.

Complement 3 is involved in changing the phenotype of human glomerular mesangial cells

Complement activation contributes to tissue injury in various forms of glomerulopathy and is characterized by deposition of complement components, which accelerates the progression of chronic renal damage. We recently reported that complement 3 (C3), a critical component of the complement system, is associated with the synthetic phenotype of vascular smooth muscle cells. It is possible that C3 stimulates mesangial cells to assume the synthetic phenotype to, in turn, induce glomerular injury and sclerosis. We investigated the role of C3 in the growth and phenotype of mesangial cells. Cultured human mesangial cells (HMCs) expressed C3 mRNA and protein, and levels were increased in response to IFN-gamma and TNF-alpha. HMCs also expressed C3a receptor mRNA and protein. Exogenous C3a stimulated DNA synthesis in HMCs in a dose-dependent manner. C3a decreased expression h-caldesmon mRNA, a marker of the contractile phenotype, and increased the expression of osteopontin, matrix Gla, and collagen type1 alpha1 (collagen IV) mRNAs, which are markers of the synthetic phenotype. C3a decreased expression of alpha-smooth muscle actin in HMCs. Small interfering RNA (siRNA) targeting C3 reduced the DNA synthesis and proliferation of HMCs, increased expression of h-caldesmon mRNA, and decreased expression of osteopontin, matrix Gla, and collagen IV mRNAs in HMCs. These results indicate that C3 causes HMCs to convert to the synthetic phenotype and stimulates growth of mesangial cells, suggesting that C3 may play an important role in phenotypic regulation of mesangial cells in renal diseases.

Assessment of linkage and association of 13 genetic loci with bone mineral density

Bone mineral density (BMD), an important risk factor for osteoporosis, is a complex trait likely affected by multiple genes. The linkage and/or association of 13 polymorphic loci of seven candidate genes (estrogen receptor alpha [ERalpha] and beta [ERbeta], calcium-sensing receptor, vitamin D receptor, collagen type 1alpha1, low-density lipoprotein [LDL] receptor-related protein 5 [LRPS], and transforming growth factor beta1) were evaluated in 177 southern Chinese pedigrees of 674 subjects, with each pedigree identified through a proband having a BMD Z score of -1.28 or less at the hip or spine. A suggestive linkage was detected between the IVS1-351A/G polymorphism of ERalpha and spine BMD, and between the 1082G/A, 1730G/A, and D14S1026 polymorphisms of ERbeta and BMD at both spine and hip. The quantitative transmission disequilibrium test (QTDT) detected total family association between 1730G/A of ERbeta and BMD at spine and hip; between D14S1026 of ERbeta and hip BMD; and between the 266A/G and 2220C/T polymorphisms of LRP5 and hip BMD. Similar total family associations were detected when only the females were analyzed. In addition, the IVS1-397T/C polymorphism of ERalpha was associated with spine BMD, and the 266A/G and 2220C/T polymorphisms of LRP5 were associated with femoral neck BMD in the females. A within-family association was detected with the IVS1-397T/C polymorphism of ERalpha, and the 266A/G and 2220C/T polymorphisms of LRP5 in the females. The effect of each polymorphism on BMD variance ranged from 1% to 4%. In conclusion, ERalpha, ERbeta and LRP5 are important candidate genes determining BMD variation, especially in females.

Isolation and partial sequencing of cDNA clones from rat incisor after Nd:YAG laser irradiation in root canal

The purpose of this study was to identify the genes expressed in odontoblast-like cells and related to osteodentin production of pulp cells in vivo during the repair process. A pulsed Nd:YAG laser was irradiated in mandibular incisors of rats at 2 W and 20 pulses/s for 5 s. We constructed a unidirectional cDNA library from the non-calcified portion of irradiated incisors, sequenced cDNA clones and classified their sequences by homology search through the GenBank database. Of the sequences determined, 26% represented sequences of new genes that were not related to any previously reported gene. 65.5% of the clones strongly matched genes and proteins in the databases. Among them, sequences found more than once were amelogenin, ameloblastin, amelin, collagen alpha1 type 1, osteonectin, nestin, enamelin and osteocalcin. Sequences detected only once were transforming growth factor beta-1 (TGF-beta1), growth hormone receptor (Ghr), glypican, enamelysin (Mmp20), amelin 2, integral membrane protein 2B, amelogenin precursor (Ame1), splicing factor 3b, insuline-like growth factor binding protein 5. This library reflects the gene expression during the dentin repair process in vivo including the odontoblast-like cell specific gene expression and osteodentin-production. Using this library, we indicate the candidate genes which might be related to odontoblast-differentiation and formation of osteodentin.

Gene expression pathways induced by axotomy and decentralization of rat superior cervical ganglion neurons

To identify genes potentially involved in remodelling synaptic connections, we induced the temporary detachment of pre- and post-synaptic elements by axotomy or denervation of rat superior cervical ganglion neurons. cDNA microarray analysis followed by stringent selection criteria allowed the identification of a panel of genes whose expression was modulated by axotomy at various time points after injury. Among these genes, 11 were validated by real-time reverse transcriptase-polymerase chain reaction on independently prepared samples after superior cervical ganglion neuron axotomy (1, 3 and 6 days) and compared with the effect of decentralization (8 h, 1 and 3 days). These genes code for extracellular matrix/space [apolipoprotein D (apoD), decorin, collagen alpha1 type I, collagen alpha1 type III] and intermediate filament (vimentin) proteins, for modulators of neurite outgrowth (thrombin receptor, plasminogen activator inhibitor-1, bone morphogenetic protein 4, annexin II and S-100-related protein, clone 42C) and for a nerve cell transcription factor (brain finger protein). Eight of these 11 genes showed significant and persistent modulations after both types of injury. Finally, protein levels of apoD were shown to increase in superior cervical ganglion after axotomy. Our results identify hitherto unrecorded genes responsive to axotomy and decentralization of superior cervical ganglion neurons, and probably involved in synapse formation, remodelling and elimination.

Ghrelin Directly Regulates Bone Formation

To clarify the role of ghrelin in bone metabolism, we examined the effect of ghrelin in vitro and in vivo. Ghrelin and its receptor, GHS-R1a, were identified in osteoblasts, and ghrelin promoted both proliferation and differentiation. Furthermore, ghrelin increased BMD in rats. Our results show that ghrelin directly affects bone formation. Ghrelin is a gut peptide involved in growth hormone (GH) secretion and energy homeostasis. Recently, it has been reported that the adipocyte-derived hormone leptin, which also regulates energy homeostasis and opposes ghrelin's actions in energy homeostasis, plays a significant role in bone metabolism. This evidence implies that ghrelin may modulate bone metabolism; however, it has not been clarified. To study the role of ghrelin in skeletal integrity, we examined its effects on bone metabolism both in vitro and in vivo. We measured the expression of ghrelin and growth hormone secretagogue receptor 1a (GHS-R1a) in rat osteoblasts using RT-PCR and immunohistochemistry (IHC). The effect of ghrelin on primary osteoblast-like cell proliferation was examined by recording changes in cell number and the level of DNA synthesis. Osteoblast differentiation markers (Runx2, collagen alpha1 type I [COLI], alkaline phosphatase [ALP], osteocalcin [OCN]) were analyzed using quantitative RT-PCR. We also examined calcium accumulation and ALP activity in osteoblast-like cells induced by ghrelin. Finally, to address the in vivo effects of ghrelin on bone metabolism, we examined the BMD of Sprague-Dawley (SD) rats and genetically GH-deficient, spontaneous dwarf rats (SDR). Ghrelin and GHS-R1a were identified in osteoblast-like cells. Ghrelin significantly increased osteoblast-like cell numbers and DNA synthesis in a dose-dependent manner. The proliferative effects of ghrelin were suppressed by [D-Lys(3)]-GHRP-6, an antagonist of GHS-R1a, in a dose-dependent manner. Furthermore, ghrelin increased the expression of osteoblast differentiation markers, ALP activity, and calcium accumulation in the matrix. Finally, ghrelin definitely increased BMD of both SD rats and SDRs. These observations show that ghrelin directly stimulates bone formation.

Bone mineral density, collagen type 1 alpha 1 genotypes and bone turnover in premenopausal women with diabetes mellitus.

Osteopenia is a recognised complication of diabetes mellitus which could be due to abnormal bone turnover or disturbances in the calcium/parathyroid hormone/vitamin D axis or both. Genetic factors also play an important part in determining bone mass although this has not been studied in diabetes. Recently a polymorphism of the collagen type 1 alpha 1 (COL1A1) gene has been shown to be associated with low bone mass in British women. To identify subjects with diabetes who may be at risk of developing osteoporosis and fractures, we analysed bone mineral density in relation to the biochemical markers of bone turnover, calcium homeostasis and the COL1A1 genotype in a group of premenopausal women with Type I (insulin-dependent) diabetes mellitus (n = 31), Type II (non-insulin dependent) diabetes mellitus (n = 21) and control subjects (n = 20). Bone mineral density was lower at the femoral neck in the subjects with Type I diabetes (p = 0.08) as were serum 25-hydroxyvitamin D compared with control subjects (p = 0.023) and this was negatively correlated with serum collagen type 1 C-terminal propeptide (r = -0.56, p < 0.001). Bone mineral density in Type II diabetes was not different from control subjects, after correction for body mass index. Bone resorption was, however, raised in the Type II diabetic subjects as reflected by the higher urinary deoxypyridinoline values (p = 0.016) and lower collagen type 1 C-terminal propeptide:deoxypyridinoline ratio (p = 0.04). In the whole group studied, subjects with the COL1A1 's' genotype had lower bone mineral density at the femoral neck (p = 0.01) which was partly attributable to a lower body mass index. Following multiple regression analysis body mass index and collagen type 1 C-terminal propeptide concentrations remained determinants of bone mass at all three sites, whereas genotype appeared to be a predictor of bone mass at the femoral neck only. We conclude that measurement of these variables could prove useful in firstly identifying those diabetic women at risk of osteoporosis and secondly guiding therapeutic intervention.