[show abstract][hide abstract] ABSTRACT: MicroRNAs (miRNAs) have been implicated in regulating diverse cellular pathways and involved in development and inflammation. This study aimed to examine six miRNAs expression during the cartilage development and identify the key miRNA which is associated with chondrogenesis.
The expression of six miRNAs in cartilage tissue during development was screened by real-time quantitative polymerase chain reaction (RT-qPCR). Rat models of bone matrix gelatin induced endochondral ossification, collagen-induced arthritis and pristane-induced arthritis were established to examine whether miR-337 is involved in chondrogenesis. Furthermore, the regulation of transforming growth factor-b type II receptor (TGFBR2) expression by miR-337 was determined with the luciferase reporter gene assay and Western blot. The expression of some specific genes relevant to cartilage tissue was tested by RT-qPCR after miR-337 mimic or inhibitor transfection.
MiR-337 expression was significantly down-regulated and almost disappeared in the maturation phases of endochondral ossification. The results of histology and RT-qPCR from three rat models showed that miR-337 is directly bound up with chondrogenesis. Furthermore, the results from the luciferase reporter gene assay and Western blot indicated that miR-337 regulated TGFBR2 expression. Our study also found that the enhancement of miR-337 may modulate the expression of cartilage-specific genes such as AGC1 in C-28/I2 chondrocytes.
We proved that miRNA-337 is associated with chondrogenesis through regulating TGFBR2 expression, and miRNA-337 can also influence cartilage-specific gene expression in chondrocytes. These findings may provide an important clue for further research in the arthritis pathogenesis and suggest a new remedy for arthritis treatment.
Osteoarthritis and Cartilage 03/2012; 20(6):593-602. · 4.26 Impact Factor
[show abstract][hide abstract] ABSTRACT: Expression profiles of microRNAs (miRNAs) can shape the repertoire of proteins expressed in development, differentiation and diseases. This study aimed to identify miRNA profile of articular cartilage at different developmental stages in rats.
Three small RNA libraries were constructed from the femoral head cartilage of Sprague-Dawley (SD) rats at postnatal day 0, day 21 and day 42 and sequenced by a deep sequencing approach. Then a bioinformatics approach was employed to distinguish genuine miRNAs from small RNAs represented in the mass sequencing data. The expression of indicated miRNAs was determined by stem-loop RT-qPCR to valuate the consistency with Solexa sequencing.
Two hundred and fifty-eight of 310 known miRNA and miRNA* genes were organized into 91 compact clusters. Two hundred and forty-six miRNAs were detected in all three small RNA libraries of rat articular cartilage. Forty-six, fifty-two and fifty-six miRNA* genes were identified from three small RNA libraries, respectively, and 86 novel miRNA candidate genes were found simultaneously. In addition, 23 known miRNAs were up-regulated (fold change ≥ 4); six were down-regulated (fold change ≤ -4) during articular cartilage development. The predicted targets of differentially expressed miRNAs were locally secreted factors and transcription factors that regulate proliferation and differentiation of chondrocytes. The same expression tendency of indicated miRNAs during articular cartilage development stages was observed by using Solexa sequencing and stem-loop RT-qPCR.
Our study provided a unique opportunity to decipher how the elaboration of the miRNA repertoire contributes to the development process of articular cartilage.
Osteoarthritis and Cartilage 07/2011; 19(10):1237-45. · 4.26 Impact Factor
[show abstract][hide abstract] ABSTRACT: Osteoporosis is a degenerative disease of the skeletal system, and its major complication is fracture that severely influences the living quality of the middle-aged and the aged. The purpose of this study was to investigate the significance of sex hormones and some biochemical indicators related to bone metabolism in the genesis and development of osteoporosis. The plasma samples were collected from 244 post-menopausal women of Xi'an urban area, and their plasma contents of testosterone, estradiol, calcitonin, osteocalcin and N-terminal propeptide of type I procollagen were detected by ELISA. The activity of tartrate-resistant acid phosphatase was determined by spectrophotometric method, and the content of nitric oxide was measured by Griess method. Bone mineral density (BMD) in lumbar vertebrae (L1-L4) and hips was measured by QDR-2000 dual energy X-ray absorptiometry. The concentrations of the biochemical indicators were compared among the three groups (normal bone mass group, osteopenia group and osteoporosis group), and Pearson correlation analysis was used to verify the correlations between the indicators and BMD. The comparison results of blood biochemical indicators of BMD-based groups showed that the plasma contents of estradiol (P = 0.006), testosterone (P = 0.038) and calcitonin (P = 0.042) decreased more significantly in the osteoporosis group, but the content of osteocalcin (P = 0.008) increased significantly in osteoporosis group than those in the other groups. The correlation analysis between BMD of different parts and the blood biochemical indicators showed that there was a significant positive correlation between estradiol and the BMD of lumber vertebra (r = 0.200, P = 0.002), femoral neck (r = 0.160, P = 0.013), and great trochanter (r = 0.204, P = 0.001). Significant positive correlations between calcitonin and BMD of lumber vertebra (r = 0.166, P = 0.018) and femoral great trochanter (r = 0.152, P = 0.041), and between testosterone and BMD of femoral great trochanter (r = 0.158, P = 0.014) were also observed. In addition, there existed significant negative correlations between osteocalcin and BMD of lumber vertebra (r = -0.220, P = 0.001), femoral neck (r = -0.259, P < 0.000), and great trochanter (r = -0.221, P = 0.001), and between the activity of tartrate-resistant acid phosphatase and BMD of femoral great trochanter (r = -0.135, P = 0.037). The partial correlation analysis also showed that there were significant correlations between estradiol (r = 0.160, P = 0.014), calcitonin (r = 0.240, P = 0.013), osteocalcin (r = -0.226, P = 0.023) and BMD when the influence of age was excluded. The Pearson correlation analysis of biochemical indicators showed there were positive correlations between the contents of testosterone and calcitonin, testosterone and osteocalcin, calcitonin and osteocalcin, calcitonin and PINP, calcitonin and NO, osteocalcin and NO, and PINP and NO, but negative correlations between the contents of testosterone and PINP, estradiol and calcitonin, estradiol and osteocalcin, and estradiol and NO. The blood contents of sex hormones and calcitonin significantly influence BMD and osteoporosis development, and the increase of osteocalcin contents could be used as a biomarker to indicate the degree of osteoporosis in post-menopausal women.
[show abstract][hide abstract] ABSTRACT: Both collagen-induced arthritis (CIA) and pristane-induced arthritis (PIA) are commonly used rat models of rheumatoid arthritis (RA). The aim of this study was systematically to compare the differences between CIA and PIA in Dark Agouti (DA) rats.
The CIA was induced by immunising DA rats intradermally with collagen type (C) and PIA was induced by injecting subcutaneously with pristane. The arthritis was evaluated macroscopically and microscopically. Nitric oxide (NO) level of plasma was determined by Griess reaction method. Plasma autoimmune antibodies, including C specific IgG antibody (anti-C IgG), cyclic citrullinated peptide specific IgG antibody (anti-CCP IgG), IgM and IgG rheumatoid factors (IgM RF and IgG RF), were detected by the enzyme-linked immunosorbent assay.
The onset of PIA rats was earlier than that of CIA rats. The involved sites of PIA rats were mostly wrist/ankle and metacarpophalangeal/metatarsophalangeal (MCP/MTP) joints while those of CIA rats were primarily distal interphalangeal (DIP) joints. NO level of plasma was increased in PIA rats, as anti-C IgG, anti-CCP IgG, IgM RF and IgG RF levels of plasma were increased in CIA rats. The kidney hyaline casts were more frequent in CIA rats than in control rats, with 9/12 in PIA group, 8/8 in CIA, and 4/8 in control, respectively.
PIA mainly affected wrist/ankle joints and MCP/MTP joints, had more severe inflammation and hardly involved other organs; while CIA mostly influenced DIP joints, had more autoimmune antibodies in plasma, and always showed hyaline casts in kidney. These findings will be useful to select the animal model of RA.
Clinical and experimental rheumatology 01/2010; 28(4):532-8. · 2.66 Impact Factor
[show abstract][hide abstract] ABSTRACT: To investigate the relationships between endothelial nitric oxide synthases (eNOS) G894T and 27 bp-variable number tandem repeat (VNTR) gene polymorphisms and osteoporosis in the postmenopausal women of Chinese Han nationality.
In the present study, 281 postmenopausal women from Xi'an urban area in West China were recruited, and divided into osteoporosis, osteopenia, and normal groups according to the diagnostic criteria of osteoporosis proposed by World Health Organization (WHO). The bone mineral density (BMD) values of lumbar vertebrae and left hips were determined by QDR-2000 dual energy X-ray absorptiometry. Blood samples were tested for plasma biochemical indicators including testosterone, estradiol, calcitonin, osteocalcin, and procollagen type I amino-terminal propeptide by enzyme-linked immunosorbent assay (ELISA), tartrate-resistant acid phosphatase by spectrophotometric method, and the content of nitric oxide by Griess method. Genome DNA was extracted from whole blood, and G894T polymorphism of eNOS gene was analyzed by using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method and 27 bp-VNTR polymorphism of eNOS gene was genotyped by PCR method. Then the relationships between genotypes and biochemical indicators, genotypes and osteoporosis, and haplotypes and osteoporosis were analyzed.
The average BMD values of the femoral neck, ward's triangle and lumbar vertebrae 1-4 (L1-L4) in the subjects with T/T genotype in eNOS G894T locus were significantly higher than those in the subjects with G/T and G/G genotypes (P<0.05). The average BMD of the femoral neck in the subjects with a/a genotype of eNOS 27 bp-VNTR locus was evidently higher than that in the subjects with b/b genotype (P<0.05). The plasma testosterone and osteocalcin concentrations in the subjects of eNOS G894T G/T genotype were evidently higher than those in the subjects of other genotypes (P<0.05); the plasma estradiol concentration in the subjects of eNOS 27 bp-VNTR a/a genotype was obviously higher than that in the subjects of b/b genotype (P<0.01). eNOS G/G homozygous frequencies in osteoporosis women, osteopenia women, and normal women were 85.37%, 76.38%, and 83.87%, respectively (P>0.05). 0% osteoporosis woman, 0.79% osteopenia women, and 3.23% normal women were eNOS a/a homozygous (P<0.05). The frequencies of eNOS 27 bp-VNTR a allele were 5.33% in the osteoporosis group, 10.24% in the osteopenia group, and 16.13% in the normal group (P<0.05, odds ratio (OR)=0.29, 95% confidence interval (CI)=0.11-0.77), suggesting that a/a genotype and a allele might have protective effects on osteoporosis. The haplotype analysis showed that G-b was 87.7% (214/244) in the osteoporosis group (P<0.05, OR=2.48, 95% CI=1.18-5.18). G-a was 5.3% (13/244) in the osteoporosis group (P<0.05, OR=0.29, 95% CI=0.11-0.77). G-b was a risk factor for osteoporosis, and G-a a protective factor.
eNOS G894T G/T genotype influenced the plasma testosterone and osteocalcin concentrations, and T/T genotype influenced BMD. eNOS 27 bp-VNTR a/a genotype increased plasma estradiol concentration to have a protective effect on osteoporosis.
Journal of Zhejiang University SCIENCE B 09/2009; 10(8):609-18. · 1.11 Impact Factor