BACKGROUND: The low survival rate of neuron cells is one of the main mechanisms of stem cell allograft, which might lead to the failure of allograft. Nuclear factor-KB (NF-kB) is one of main transcription factors for cell signaling transduction and participates in call proliferation and differentiation.OBJECTIVE: To study the differentiation of muscle-derived stem cells (MDSCs) into neuron-like cells induced by ciliary neurotrophic factor (CNTF) and Compound Salvia Miltiorrhiza Injection in vitro, and the expression of NF- kB.DESIGN, TIME AND SETTING: The experiment at cell molecular level was performed at the Oral Science Laboratory of the Second Affiliated Hospital of Liaoning Medical College from March to May 2008.MATERIALS: A total of 10 Sprague Dawley neonatal rats aged 7 days were supplied by Experimental Animal Center, Liaoning Medical University. CNTF (Sigma, USA) and Compound Salvia Miltiorrhiza Injection (Chiatai Qingchunbao Pharmacantical Co.,Ltd., China) were used in this study.METHODS: Rat MDSCs were harvested in vitro, pudfied by differential adherence and enzyme digestion, and incubated in 6-well plate. Samples in the induction group were incubated in DMEM containing CNTF for 24 hours. The medium was changed.Subsequently, samples were dnsed three times, and then incubated in serum-free DMEM supplemented with Compound Salvia Miltiorrhiza Injection for 5 hours. Samples in the control group were treated with serum-free DMEM.MAIN OUTCOME MEASURES: Neurofilament protein and NF-KB inhibitor protein expression were detected using reverse transcription-polymerase chain reaction (RT-PCR) and Westem blotting.RESULTS: No neurofilament protein expression was found in MDSCs before induction, and neurofilament protein-positive MDSCs were detected following induction. Results of gel electrophoresis and Westam blot showed that no significant differences in NF-kB inhibitor protein expression were determined in the control group, and NF-kB inhibitor protein expression was significantly decreased in the induction group after induction.CONCLUSION: CNTF and Compound Salvia Miltiorrhiza Injection could inhibit the activation of NF-kB and induce the differentiation of MDSCs into neuron-like cells.Zeng XY, Wang W, Sun L, Zhang L, Zeng LD.Differentiation of muscle-derived stem cells into neuron-like cells induced by ciliary neurotrophic factor and Compound Salvia Miltiorrhiza Injection in vitro.Zhongguo Zuzhi Gongcheng Yanjiu yu Linchuang Kangfu.2009;13(27): 5336-5340. [http://www.crter.cn http://en.zglckf.com]

[ ABSTRACT] AIM: To investigate the underlying genetic changes of a Chinese patient with infantile malignant osteopetrosis ( IMO) .IMO is a monogenic disease, mostly caused by mutations of TCIRG1 and CLCN7 genes.The former is believed a homozygous gene and only cause the disease in homozygous or compound heterozygous status.However, it has been reported that heterozygous mutations also cause the disease in 6 non-Chinese cases.METHODS:Genomic DNA was extracted from peripheral blood of the patient and his parents.All exons and splice sites of TCIRG1 and CLCN7 genes were amplified by PCR followed by Sanger sequencing.Mutation detection in the 2 genes was also investigated in the parents. Haplotypes were constructed by variations obtained in mutation detection and microsatillites flanking TCIRG1 gene in the family by Cyrillic.Chromosomal microarray analysis ( CMA) was performed to detect copy number variations ( CNV) of the patient and his mother.RESULTS:A novel mutation c.449_452delAGAG ( p.Gln149Glnfs16) was detected in the pa-tient.This mutation truncated 666 amino acids at the C terminal of the V-ATPase 116 kD isoform a3 protein.It wiped out the entire ATPase V0 complex and was predicted to result in total loss of protein function.This mutation was also detected in the patient’ s father.No pathogenic mutation was detected in CLCN7 gene.CMA did not reveal any CNV involving TCIRG1 or CLCN7 gene.CONCLUSION:We reported a novel heterozygous mutation of TCIRG1 gene causing IMO.This represents the first IMO case in China caused by heterozygous TCIRG1 gene mutation.

OBJECTIVEThis study aimed to evaluate the biological characteristics of a human specifically targeted antimi- crobial peptide C16LL-37 against Streptococcus mutans (S. mutans).

METHODSIn this study, an antimicrobial peptide LL-37, a peptide derived from CSP(C16) (S. mutans competence stimulating peptide), and recombinant peptide C16LL-37 were synthesized by Fmoc-chemistry-based strategy. The selectivity and antibacterial activity of C16LL-37 were identified by the colony counting method on microbial culture plates. After treatment of C16LL-37 at 32 µmol · L⁻¹, the morphological changes in S. mutans were observed by using scanning electron microscopy (SEM). In addition, enzyme-linked immunosorbent assay was used to evaluate the hemolytic activity and antibacterial activity of C16LL-37 under different conditions.

RESULTS1) The minimum inhibitory concentration of C16LL-37 was 16 µmol · L⁻¹, and the minimum bactericidal concentration was 64 μmol ·L⁻¹. 2) The survival rate of S. mutans was 3.46% after C16LL-37 treatment at 64 µmo-L⁻¹ for 30 min, whereas it was 0% at 64 µmol · L⁻¹ for 60 min. The survival rates of four other kinds of bacteria were more than 60% at any time (P < 0.05). 3) The morphological change in S. mutans was observed after C16LL-37 treatment at 32 µmol · L⁻¹ by using SEM. S. mutans presented an irregular shape, rough surface, and evident splitting. 4) The hemolysis rate of C16LL-37 (≤ 64 µmol · L⁻¹) was less than 0.33%. 5) This study showed no significant in- fluence on the antibacterial activity of C16LL-37 under different conditions, such as temperature, pH, salinity, and trypsin at low concentration (P > 0.05).

CONCLUSIONC16LL-37 exhibited obvious specificity for S. mutans, strong antibacterial activity, low toxicity, and high stability. Thus, C16LL-37 has good potential in caries research and clinical application.

Anti-Infective Agents ; pharmacology ; Antimicrobial Cationic Peptides ; pharmacology ; Bacterial Proteins ; Dental Caries ; Enzyme-Linked Immunosorbent Assay ; Humans ; Microbial Sensitivity Tests ; Microscopy, Electron, Scanning ; Peptides ; Streptococcus mutans ; drug effects

Cleidocranial dysplasia (CCD) is a rare congenital disorder, typically characterized by persistently open skull sutures, aplastic or hypoplastic clavicles, and supernumerary teeth. Mutations in the gene encoding the runt-related transcription factor 2 (RUNX2) protein are responsible for approximately two thirds of CCD patients. We report a 20-year-old CCD patient presenting not only with typical skeletal changes, but also complex dental anomalies. A previously undiagnosed odontoma, 14 supernumerary teeth, a cystic lesion, and previously unreported fused primary teeth were discovered on cone-beam computed tomography (CBCT) scans. Mutation analysis identified the causal c.578G>A (p.R193Q) mutation in the RUNX2 gene. At 20 years of age, the patient had already missed the optimal period for dental intervention. This report describes the complex dental anomalies in a belatedly diagnosed CCD patient, and emphasizes the significance of CBCT assessment for the detection of dental anomalies and the importance of early treatment to achieve good outcomes.
Clavicle ; Cleidocranial Dysplasia* ; Cone-Beam Computed Tomography ; Congenital, Hereditary, and Neonatal Diseases and Abnormalities ; Humans ; Odontoma ; Skull ; Sutures ; Tooth, Deciduous ; Tooth, Supernumerary ; Transcription Factors ; Young Adult

OBJECTIVETo construct a cell line of oral mucosa epithelial cells that stably express human telomerase reverse transcriptase (hTERT) by lentiviral vectors, approaches for the establishment of stable and efficient immortalized oral mucosa epithelial cell lines were explored.

METHODSWhole RNA was extracted from 293T cells. The hTERT gene was amplified by polymerase chain reaction (PCR) and cloned into the lentiviral vector as pLVX-puro-hTERT. The lentivirus particles were successfully packaged and used to infect primary oral epithelial cells. The positive cell clones were selected by puromycin. Finally, the expression of hTERT was examined by real-time fluorescent quantitative PCR (qRT-PCR) and Western blot analysis.

RESULTSThe sequencing results confirmed the construction of the recombinant lentivirus pLVX-puro-hTERT. The morphology of infected cells was similar to that of normal oral mucosal epithelial cells, with a cobble stone-like appearance. The qRT-PCR and Western blot results showed that hTERT was overexpressed in infected cells compared with the normal group (P<0.05).

CONCLUSIONSThe oral epithelial cell line with stable expression of hTERT was successfully established by the lentivirus, which provides an experimental basis for the establishment of a highly efficient and stable oral epithelial immortalized cell line.

HEK293 Cells ; Humans ; Lentivirus ; Mouth ; Mucous Membrane ; Real-Time Polymerase Chain Reaction ; Telomerase