Anti-inflammatory and repaired effects of non-steroidal anti-inflammatory drugs on human dental pulp cells / 北京大学学报(医学版)

OBJECTIVE@#To study the effects of non-steroidal anti-inflammatory drugs (NSAIDs) on anti-inflammation and repair of human dental pulp cells (hDPCs).@*METHODS@#Primary hDPCs from the freshly extracted human third molars were cultured and passaged in vitro, and the following experiments were performed using the 4th-6th generations of hDPCs. HDPCs were cultured in Dulbecco's modified eagle medium (DMEM) containing 1 mg/L lipopolysaccharide (LPS) to obtain LPS irritated hDPCs (LPS-hDPCs), which served as the inflammatory positive group. LPS-hDPCs in the experimental group were cultured in DMEM containing different concentrations (1, 10, and 100 μmol/L) of NSAIDs (aspirin or meloxicam). HDPCs cultured in DMEM were used as the negative control group. The effects of NSAIDs on the proliferation of hDPCs were assessed on the 1st, 3rd, 5th, and 7th day by MTT assay. The effects of NSAIDs on the expression of inflammation related genes interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) of LPS-hDPCs were detected at the 6th hour by real-time PCR. The expression of differentiation related markers dentin matrix protein-1 (DMP-1) and dentin sialophosphoprotein (DSPP) were detected on the 7th day by real-time PCR. The effects of NSAIDs on the mineralization of LPS-hDPCs were assesd on the 14th day by alizarin red staining. Calcium mineralized nodules were semi-quantitatively determined by cetyl pyridine chloride.@*RESULTS@#MTT assay showed that 1-100 μmol/L aspirin or meloxicam significantly promoted the proliferation of hDPC in a concentration dependent manner (P<0.05). Real-time PCR showed that 1-100 μmol/L meloxicam or 100 μmol/L aspirin down-regulated significantly the mRNA expression of TNF-α and IL-6 of LPS-hDPCs (P<0.05), and 100 μmol/L meloxicam down-regulated IL-6 and TNF-α more significantly than 100 μmol/L aspirin of LPS-hDPCs (P<0.05). Real-time PCR showed that 100 μmol/L meloxicam up-regulated the mRNA expression of DMP-1 and DSPP of LPS-hDPCs significantly (P<0.05). Alizarin red staining showed the meloxicam at the concentration of 100 μmol/L significantly promoted the mineralization of LPS-hDPCs (P<0.05).@*CONCLUSION@#In this study, meloxicam promoted the proliferation of hDPCs, inhibited the inflammatory reaction and promoted differentiation and mineralization of hDPCs under LPS irritation. The present results suggest that meloxicam may play a role in anti-inflammation and repair of pulp inflammation.

Semaphorin 3A-hypoxia inducible factor 1 subunit alpha co-overexpression enhances the osteogenic differentiation of induced pluripotent stem cells-derived mesenchymal stem cells in vitro / 中华医学杂志(英文版)

BACKGROUND@#Mesenchymal stem or stromal cells (MSCs) derived from the induced pluripotent stem cells (iPSCs) have uniform biological activity, which makes the clinical application of MSCs in bone repair possible. Culturing the iPSC-MSCs onto osteoconductive materials is a promising tissue engineering-based strategy in bone regeneration. The aim of this work was to evaluate the effects of semaphorin 3A (Sema3A) and hypoxia inducible factor 1 subunit alpha (HIF1α) co-overexpression on the survival and osteogenic differentiation of iPSC-MSCs.@*METHODS@#Sema3A and HIF1α were linked together with the three (GGGGS; G, glycine; S, serine) peptide fragment, and their co-expression in iPSC-MSCs was mediated by a lentiviral vector. The fusion protein retained the immune reactivity for both Sema3A and HIF1α as determined with Western blotting. iPSC-MSCs were infected with overexpression lentivirus (oeLenti) as negative control, oeLenti-Sema3A, oeLenti-HIF1α or oeLenti-Sema3A-HIF1α lentiviruses.@*RESULTS@#Sema3A overexpression alone promoted the osteogenic differentiation of iPSC-MSCs (the activity and/or expression of osteoblast markers, such as alkaline phosphatase, osteopontin, and osteocalcin, were upregulated), and suppressed cell survival. The Sema3A-HIF1α fusion protein showed a comparable osteoconductive effect to that of Sema3A without reducing cell survival. We further seeded iPSC-MSCs modified by SemaA-HIF1α overexpression onto hydroxyapatite (HA) scaffolds, and evaluated their growth and differentiation on this three-dimensional material. Additional data indicated that, as compared to iPSC-MSCs cultured in ordinary two-dimensional dishes, cells cultured in HA scaffolds grew (blank vs. HA scaffolds: 0.83 vs. 1.39 for survival) and differentiated better (blank vs. HA scaffolds: 11.29 vs. 16.62 for alkaline phosphatase activity).@*CONCLUSION@#Modifying iPSC-MSCs with pro-osteogenic (Sema3A) and pro-survival (HIF1α) factors may represent a promising strategy to optimize tissue engineering-based strategy in bone repair.

Claudin-3 expression in colorectal carcinoma and its significance / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the expression of claudin-3 in colorectal carcinoma and its association with the occurrence, progression and prognosis of colorectal cancer.</p><p><b>METHODS</b>Forty surgical specimens of colorectal carcinoma and 22 adjacent normal tissues resected between October, 2010 and January, 2013 at Nanfang Hospital were examined for claudin-3 expression using immunohistochemistry, which was analyzed in association with the clinicopathological parameters and the survival of the patients.</p><p><b>RESULTS</b>Claudin-3 was expressed mainly on the cell membrane, and its positivity rate was significantly higher in cancer tissues than in normal tissues (92.50% vs 59.09%, P<0.05). In 13 cases claudin-3 expression was detected in both the cancer tissues and adjacent normal tissues with average expression scores of 4.538 and 3.269, respectively (P<0.05). In the cancer tissues, the strongly positive expression rate was significantly higher in poorly differentiated tissues (85.71%) than in well (21.43%) and moderately (36.48%) differentiated tissues (P<0.05), and was higher in cases with lymph node metastasis than in those without (61.11% vs 22.72%, P<0.05). The strongly positive expression rate of claudin-3 was not correlated with the patients'age, gender, tumor location or tumor size (P>0.05). Of the 33 cancer patients followed up, 14 had a postoperative survival time no longer than 3 years and 19 had longer survival time, and their average expression scores differed significantly (4.50 vs 3.526, P<0.05).</p><p><b>CONCLUSION</b>Claudin-3 is over-expressed in colorectal cancer tissues, and its high expression may promote the occurrence and progression of colorectal cancer. Claudin-3 may serve as a molecular biomarker for early diagnosis and prognostic evaluation.</p>

Functional Annotation of Metastasis-associated MicroRNAs of Melanoma: A Meta-analysis of Expression Profiles / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Melanoma is a type of cancer that develops from the pigment-containing cells. Until now, its pathological mechanisms remain largely unknown. The aim of this study was to identify metastasis-related microRNA (miRNAs) and gain an understanding of the biological functions in the metastasis of melanoma.</p><p><b>METHODS</b>We searched the PubMed and Gene Expression Omnibus database to collect miRNA expression profiling datasets about melanoma, with key words of "melanoma", "miRNA", "microarray", and "gene expression profiling". Only the original experimental works published before June 2016 for analyzing the metastasis of melanoma were retained, other nonhuman studies, reviews, and meta-analyses were removed. We performed a meta-analysis to explore the differentially expressed miRNA between metastatic and nonmetastatic samples. Moreover, we predicted target genes of the miRNAs to study their biological roles for these miRNAs.</p><p><b>RESULTS</b>We identified a total of 63 significantly differentially expressed miRNAs by meta-analysis of the melanoma expression profiling data. The regulatory network constructed by using these miRNAs and the predicted targets identified several key genes involved in the metastasis of melanoma. Functional annotation of these genes indicated that they are mainly enriched in some biological pathways such as mitogen-activated protein kinase signaling pathway, cell junction, and focal adhesion.</p><p><b>CONCLUSIONS</b>By collecting the miRNA expression datasets from different platforms, multiple biological markers were identified for the metastasis of melanoma. This study provided novel insights into the molecular mechanisms underlying this disease, thereby aiding the diagnosis and treatment of the disease.</p>

Relationship between metabolic syndrome and bone mineral density in the elderly aged over 60 years in Shanghai / 第二军医大学学报

Objective To investigate the relationship between metabolic syndrome (MS) and bone mineral density(BMD) in the aged. Methods 1002 cases were divided into MS group and non-MS(NMS)group according to with or without MS. Height, weight, waist circumference and blood pressure were measured, body mass index(BMI) were calculated, total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), low density lipoprotein cholesterol (LDL-C), fasting blood glucose (FBG) were tested. The calcaneal bone mineral density (BMD) was measured by the quantitative ultrasonography. To find the differences of BMD between subjects with or without MS by using the covariance analysis and to analyze the risk factors associated with BMD. Results Compared to NMS group, the values of BMI, waist circumference, TC, TG, FPG were significantly increased in MS group (P<0.05 P<0.01).Those with MS had higher BMD than those without after adjustment for age and BMI (P<0.01). There was no significant difference in BMD among patients with different number of the components of the metabolic syndrome (P<0.05). However，the difference were significant after adjustment for age and BMI (P<0.05).The multiple stepwise regression analysis demonstrated that BMD were correlated with age, waist circumference and HDL-C (P<0.05).Conclusion .The BMD is higher in aged MS patients than that in non-MS subjects.

Functional annotation of the microRNA-mediated network in gigantomastia by integrating microRNA and mRNA expression profiling / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Gigantomastia is the overdevelopment of the female mammary gland, and it causes great physiological and psychological burdens to patients. A better understanding of the molecular mechanisms involved in gigantomastia is needed to develop less invasive and more effective medical treatments. MicroRNA (miRNA) is a class of small noncoding RNAs that play an important regulatory role at the post-transcriptional level. These miRNAs are known to be involved in many diseases, including breast cancer; however, the relationship between miRNA and gigantomastia is largely unknown.</p><p><b>METHODS</b>Whole genome-wide expression of miRNA and mRNA in gigantomastia was detected using microarray and functional annotation was performed based on the altered expression of miRNAs and mRNAs.</p><p><b>RESULTS</b>We found many miRNAs and mRNAs to be significantly differentially expressed in gigantomastia compared with normal breast tissues. Functional annotation analysis indicated that APK, Wnt, and Neurotrophin signaling pathways may participate in gigantomastia.</p><p><b>CONCLUSION</b>This study addresses the need for better diagnosis and treatment of gigantomastia by providing new insight into the molecular mechanisms underlying this disease.</p>

P1 gene of Mycoplasma pneumoniae in clinical isolates collected in Beijing in 2010 and relationship between genotyping and macrolide resistance / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Mycoplasma pneumoniae is a common pathogen that caused community-acquired pneumonia (CAP). P1 protein served as major adhesion and immunodominant protein in Mycoplasma pneumoniae, but little about P1 gene was learned and the relationship between P1 genotype and macrolide resistance has yet to be explored.</p><p><b>METHODS</b>The DNA sequence of the entire P1 gene from 35 strains isolated from clinical specimens collected in Beijing, China, in 2010 was determined. The resulting sequences were checked for known macrolide resistance mutations, such as A2063G, A2064G, C2617G in domain V of 23S rRNA. Antibiotic susceptibility test was done to further identify macrolide resistant strains.</p><p><b>RESULTS</b>Thirty-four clinical strains were type 1, and were identical to type 1 reference strain MP129. Only one clinical strain, MpYYM22, was type 2, and proved to be variant 2c. One synonymous point mutation in the P1 type 1 gene from two isolates was identified relative to the MP129 P1 sequence at nucleotide position (nt) 552 (C>A), while another two isolates had missense mutations at nt 2504 (G>A). This point mutation caused an amino acid change from glycine to glutamic acid. An AGT tri-nucleotide variable-number tandem repeat (VNTR), coding for serine and repeating 6-11 times, up to 15-16 times, was found in the region between the RepMP4 and RepMP2/3 elements in the 35 isolates examined. All 35 clinical strains, including MpYYM22, demonstrated macrolide resistance with the range of minimum inhibitory concentration (MIC) of erythromycin from 64 to 256 µg/ml, having an A2063G transition in domain V of the 23S rRNA gene.</p><p><b>CONCLUSIONS</b>P1 type 1 was the dominant type of Mycoplasma pneumoniae in Beijing in 2010, although variant 2c strains were present. More samples are needed to determine whether there is a relationship between the P1 genotype and macrolide resistance, as the 35 strains examined did not allow a conclusive result. However, the AGT tri-nucleotide VNTR may be a more informative locus for multi-locus VNTR analysis.</p>

Logistic regression analysis of damp-heat and cold-damp impeding syndrome of rheumatoid arthritis: a perspective in Chinese medicine / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate a method for quantitative differential diagnosis of damp-heat and cold-damp impeding syndrome of rheumatoid arthritis (RA) in Chinese medicine (CM).</p><p><b>METHODS</b>Laboratory parameters were collected from 306 patients with RA. The clinical symptoms and laboratory parameters were compared between patients with these two syndromes (158 with RA of damp-heat impeding syndrome, and 148 with RA of cold-damp impeding syndrome), and a regression equation was established to facilitate discrimination of the two RA syndromes.</p><p><b>RESULTS</b>There were significant differences in disease activity score in 28 joints [DAS28 (4)], erythrocyte sedimentation rate (ESR), white blood cell count (WBC), C-reactive protein (CRP), platelet count (PLT), albumin (ALB) and globulin (GLB) between the two syndrome of RA (P<0.05). Logistic regression analysis showed that the parameters ESR, WBC, CRP, joint pyrexia, joint cold, thirst, sweating, aversion to wind and cold, and cold extremities were statistically useful to discriminate damp-heat from cold-damp impeding syndrome. The regression equation was as follows: P=1/{1+exp[-(3.0-0.021X (1)-0.196X (2)-0.163X (3)-1.559X (4)+1.504X (5)-0.927X (6)-1.039X (7)+1.070X (8)+1.330X (9))]}. The independent variables X (1)-X (9) were ESR, WBC, CRP, hot joint, cold joint, thirst, sweating, aversion to wind and cold, and cold limbs. A P value > 0.5 signified cold-damp impeding syndrome, and a P value < 0.5 signified damp-heat impeding syndrome. The accuracy was 90.2%.</p><p><b>CONCLUSION</b>The regression equation may be useful for discriminating damp-heat from cold-damp impeding syndrome of RA.</p>

The effects of extremely low frequency electromagnetic field exposure on the pH of the adult male semen and the motoricity parameters of spermatozoa in vitro / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To study the effects of 50-Hz extremely low frequency electromagnetic field (ELF-EMF) exposure on the pH of the adult male semen and the motoricity and motoricity parameters of spermatozoa.</p><p><b>METHODS</b>Healthy adult male fresh semen was exposed to a 50-Hz EMF at 0.4 mT for 15, 30 and 60 min, respectively. The pH value of the semen, the motoricity and motoricity parameter of spermatozoa were detected and recorded in real time using the WLJY-9000 pattern chromatic color spermatozoa quality detection system.</p><p><b>RESULTS</b>Compared with parallel control group, the exposure of adult male fresh semen to a 50-Hz EMF at 0.4 mT for 15 min or 60 min could decrease significantly the motoricity (spermatozoa with a + b lever) and the activity ratio (spermatozoa with a + b + c lever)(P < 0.01). However, there were no significant differences of motoricity and the activity ratio between exposure group and control group (P > 0.05), and after exposure to a 50-Hz. EMF for 30 min the motoricity and the activity ratio of exposure group were inhibited, as compared with control group (P < 0.01 or P < 0.05). The pH value of the semen was not obvious changed (P > 0.05) when semen was exposed to a 50-Hz EMF of 0.4 mT for 15, 30 and 60 min.</p><p><b>CONCLUSION</b>In present experiment, it is suggested that the exposure of adult male fresh semen to a 50-Hz EMF in vitro could inhibit the motoricity and the activity ratio, but not affect the pH value of the semen within 60 min.</p>

Epigenetic regulation of Wnt signaling pathway gene SRY-related HMG-box 17 in papillary thyroid carcinoma / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>SRY-related HMG-box 17 (SOX17) encodes a member of the SOX (SRY-related HMG-box) family of transcription factors involved in the regulation of embryonic development and in the determination of the cell fate. Recently, it was considered as a tumor suppressor gene to inhibit canonical Wnt/β-catenin signaling pathway in several malignancies. However, the function of SOX17 in thyroid cancer was unknown. Therefore, we investigated the epigenetic changes and the function of SOX17 in thyroid cancer.</p><p><b>METHODS</b>The methylation status of the promoter region of SOX17 was detected using methylation-specific PCR in 63 papillary thyroid carcinoma (PTC) tissue, 10 normal thyroid tissue, and two thyroid cancer cell lines. Semi-quantitative RT-PCR was used to assess mRNA expression of SOX17 before and after 5-aza-2'-deoxycytidine treatment in thyroid cancer cell lines. Expression of SOX17 and β-catenin were detected by immunohistochemistry in PTC and adjacent tissue. Luciferase reporter assay, colony formation, transfection, and Western blotting were employed to analyze the effect of SOX17 on thyroid cancer cell proliferation and the function of SOX17 in the Wnt signal pathway.</p><p><b>RESULTS</b>Loss of SOX17 expression was correlated to the promoter region hypermethylation in thyroid cancer cell lines. Re-expression of SOX17 was found in TPC-1 cell line after 5-aza-2'-deoxycytidine treatment. In primary thyroid cancer, 60.3% (38/63) were methylated and 39.7% (25/63) unmethylated. But no methylation was found in noncancerous thyroid tissues. Methylation of SOX17 was associated reversely with β-catenin expression in the cytoplasm or nucleus significantly in the PTC (P < 0.05). Colony formation was inhibited by re-expression of SOX17 in TPC-1 cells. SOX17 suppressed the Wnt signaling pathway and the HMG domain was essential for this effect.</p><p><b>CONCLUSIONS</b>SOX17 was frequently methylated in human PTC. Loss of SOX17 expression was induced by promoter region hypermethylation. SOX17 inhibited thyroid cancer proliferation. Methylation of SOX17 activated the Wnt signaling pathway in human thyroid cancer.</p>

Involvement of LMO4 in tumorigenesis associated epithelial-mesenchymal transition / 浙江大学学报·医学版

LMO4 is a novel member of the LIM-only (LMO) subfamily of LIM domain-containing transcription factors, so named because they are composed almost entirely of two tandem LIM domains. This subgroup of LIM proteins has 4 members: LMO-1, LMO-2, LMO-3 and LMO-4. They all play important roles in the normal mammalian development, functioning as an important regulator of cell proliferation. LMO4 is highly expressed in the epithelial compartments at locations of active epithelial-mesenchymal interactions, and can interact with some signaling pathways involved in epithelial-mesenchymal signaling. Thus the disregulation of LMO4 expression may be involved in tumorigenesis. In this paper, we will at first expound LMO4 in detail, based on which the possible mechanisms for its interaction with TGF-β signaling and the roles of this cross-talk between them in the vital process of cell will be introduced. All of those will add to our understanding of tumorigenesis and contribute to the search of new targets for the treatment of cancer.

Preliminary analysis of cerebrospinal fluid proteome in patients with neurocysticercosis / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Neurocysticercosis is the infection of the nervous system by the larvae of Taenia solium (T. solium). Despite continuous effort, the experimental diagnosis of neurocysticercosis remains unresolved. Since the cerebrospinal fluid (CSF) contacts with the brain, dynamic information about pathological processes of the brain is likely to be reflected in CSF. Therefore, CSF may serve as a rich source of putative biomarkers related to neurocysticercosis. Comparative proteomic analysis of CSF of neurocysticercosis patients and control subjects may find differentially expressed proteins.</p><p><b>METHODS</b>Two-dimensional difference in gel electrophoresis (2D-DIGE) was used to investigate differentially expressed proteins in CSF of patients with neurocysticercosis by comparing the protein profile of CSF from neurocysticercosis patients with that from control subjects. The differentially expressed spots/proteins were recognized with matrix-assisted laser desorption/ionization-time of flight-time of flight (MALDI-TOF-TOF) mass spectrometry.</p><p><b>RESULTS</b>Forty-four enzyme digested peptides were obtained from 4 neurocysticercotic patients. Twenty-three were identified through search of the NCBI protein database with Mascot software, showing 19 up-expressed and 4 down-expressed. Of these proteins, 26S proteosome related to ATP- and ubiquitin-dependent degradation of proteins and lipocalin type prostaglandin D synthase involved in PGD2-synthesis and extracellular transporter activities were up-expressed, while transferrin related to iron metabolism within the brain was down-expressed.</p><p><b>CONCLUSIONS</b>This study established the proteomic profile of pooled CSF from 4 patients with neurocysticercosis, suggesting the potential value of proteomic analysis for the study of candidate biomarkers involved in the diagnosis or pathogenesis of neurocysticercosis.</p>

Expression, purification and antibody preparation of recombinant human SM22alpha / 中国应用生理学杂志

<p><b>AIM</b>The recombinant human smooth muscle 22 alpha (SM22alpha) was expressed by using Pichia pastoris.</p><p><b>METHODS</b>Using pGEM3z-SM22alpha as the template, SM22alpha coding region was amplified by PCR, and was inserted the expression vector pPIC9. Then the recombinant plasmid pPIC9-SM22alpha was transfected into Pichia pastoris. The products induced by methanol were precipitated by ammonium sulfate, then CM-cellulose chromatography was performed for SM22alpha. Polyclonal antibody against SM22alpha was produced by immunizing a rabbit with purified recombinant SM22alpha.</p><p><b>RESULTS</b>The positive clone with SM22alpha got high output at 84 hours after induction by methanol. The SM22alpha prepared by ammonium sulfate fractionation and chromatographic separation showed a single band whose apparent molecular weight was 22 kD on SDS-PAGE. Polyclonal antibody against SM22alpha could detect the SM22alpha expression in human or rat vascular walls.</p><p><b>CONCLUSION</b>High-level expression of SM22alpha is successfully achieved in Pichia pastoris. Antibody against SM22alpha can be used to explore the function of SM22alpha.</p>