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Objective To compare in-vitro percutaneous absorption and pharmacodynamic actions of anti-inflammation and inhibiting delayed-type hypersensitivity of Chinese herbal compound cremor for eczema (CHCCE) with different mass concentrations of synthetic borneol. Methods By adopting modified Franz diffusion device add with isolated BALB/cnude mice skin as a barrier, in vitro percutaneous absorption effectiveness of CHCCE with different mass concentrations of borneol was compared by in vitro percutaneous test after the content of matrine was determined with high performance liquid chromatography(HPLC). Meanwhile, the effects of CHCCE with different mass concentrations of synthetic borneol on reducing dimethylbenzene-induced auricular edema and suppressing delayed-type hypersensitivity induced by 2,4-dinitrochlorobenzene(DNCB) in mice were compared. Results Cumulative permeation amount of matrine in CHCCE with synthetic borneol was higher than that in CHCCE without synthetic borneol 2～ 48 h after administration (P 0.05) among CHCCE groups with different mass concentrations of synthetic borneol after 48 h. In vitro percutaneous absorption behavior of matrine arrived to the steady state and the cumulative permeation amount of matrine presented a decreasing trend in all medication groups 12 h after administration. Within 12 h of the medication, the permeation rate of CHCCE with different mass concentrations of borneol was in the sequence of 3％ borneol > 1％ borneol > 2％ borneol > 0.5％ borneol > no borneol. The content of matrine was decreased with the increase of mass concentration of synthetic borneol after 12 h. The results of pharmacodynamic actions of CHCCE showed that compared with the blank control group, CHCCE with 1％, 3％ synthetic borneol could significantly suppress the acute inflammation induced by dimethylbenzene and inhibit contact dermatitis induced by dinitrochlorobenzene (DNCB) in mice(P 0.05). Conclusion CHCCE with 1％ synthetic borneol has good effects on in vitro transdermal absorption, and can suppress inflammation and delayed-type hypersensitivity effectively.
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Background: Anal fissure is one of the most common anal-rectum diseases, and approximately 10 percent patients with chronic anal fissure ultimately receive surgery. Relieving postoperative pain and protecting functions of the sphincter are central issues for coloproctologists. Objective: To evaluate the efficacy and safety of anoplasty in the treatment of chronic anal fissures. Design, setting, participants and interventions: In this prospective, multicenter, randomized controlled trial, 120 adult patients with chronic anal fissure were referred from Department of Coloproctology of Yueyang Hospital of Integrated Traditional Chinese Medicine Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai Ninth People's Hospital Affiliated to Shanghai Jiao Tong University School of Medicine and Shanghai Municipal Hospital of Traditional Chinese Medicine. The patients were enrolled from January 2009 to April 2010 and randomly divided into study (mucosa advancement flap anoplasty, abbreviated as anoplasty) group and control (fissurectomy) group. The two groups were assessed separately, and the main outcome measures were observed for 2 weeks, with a short-term follow-up for 6 weeks. Main outcome measures: Degree of pain, haemorrhage and anal canal pressure were observed and recorded preoperatively, and on the third day, the fourteenth day and the sixth week postoperatively. The wound healing time was also recorded. Surgical complications of the two groups were recorded and compared on the third day and the sixth week postoperatively. The curative effects associated with the surgery were analyzed on the fourteenth day and the sixth week after surgery and the therapeutic results were evaluated. Results: Three patients were dropped out due to the early discharge from hospital and losing connection (1 in study group and 2 in control group). Overall the surgery showed that the anoplasty group had better results than the fissurectomy group in the curative effect on the sixth week after operation (P0.05). There were no significant differences in relieving the anal canal pressure (P>0.05) and the surgical complications (dysuria, edema of anal margin, fever, infection, anal incontinence and anal deformation) between the two groups (P>0.05). None of the patients suffered postoperative complications by the sixth week after operation. Furthermore, there was no recurrence in either of the two groups at six weeks after operation. Conclusion: The results indicate that anoplasty for chronic anal fissures has advantages such as better therapeutic effects, less postoperative pain, a shorter healing time and no incidence of anal incontinence.
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To observe the effects of Fuhuang Shengji Yuchuang (FHSJYC) Ointment, a compound traditional Chinese herbal medicine, on the expressions of types I and III collagens in granulation tissue of wound in rats with diabetes.
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OBJECTIVE: To study the effects of resolving stagnation and promoting granulation therapy on expressions of Bax and Bcl-2 in granulation tissue of diabetic rats during wound healing. METHODS: Seventy-two male SD diabetic rats with full-thickness skin lesion were randomly divided into 3 groups: SJHYR 1-treated group, SJHYR 2-treated group and normal saline (NS) control group. SJHYR 1 was prepared with Shengji Recipe (SJR, a compound traditional Chinese herbal medicine for promoting granulation) and Huayu Recipe (HYR, a compound traditional Chinese herbal medicine for resolving stagnation) at a ratio of 1:2, while SJHYR 2 was prepared with SJR and HYR at a ratio of 1:1. Immunohistochemical method was used to assess Bax and Bcl-2 protein levels in granulation tissue. RESULTS: SJHYR 1 could accelerate wound healing as compared with SJHYR 2 and NS (P<0.05). On the third day in experiment, Bax and Bcl-2 proteins were not found in any groups, but on the seventh and eleventh day in experiment, Bax and Bcl-2 proteins in SJHYR 1-treated group were much higher than those in the other two groups (P<0.05). CONCLUSION: SJR and HYR in different ratios may all have a role in regulating Bax and Bcl-2 expression in granulation tissue of diabetic rats during wound healing.
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OBJECTIVE: To study the effects of Shengji Huayu Recipe (a traditional Chinese medicine compound recipe for resolving stagnation and promoting granulation) and its decomposed formulas (Huayu Recipe for resolving stagnation and Shengji Recipe for promoting granulation) on the synthesis of collagen types I and III in granulation tissue of rats in early wound healing. METHODS: Twenty-four male Sprague-Dawley (SD) rats with full-thickness skin lesion were randomized into 4 groups: Shengji Huayu Recipe-treated group, Shengji Recipe-treated group, Huayu Recipe-treated group and untreated group. Collagen types I and III in granulation tissue of the rats were tested with immunohistochemical methods and image analysis. RESULTS: On the third day of wound healing, collagen I of the rats in both Shengji Huayu Recipe-treated group and Shengji Recipe-treated group was higher than that in the untreated group, and collagen I of the rats in Huayu Recipe-treated group was lower than that in the untreated group (P<0.05). Collagen III of the rats in the three treated groups were lower than that in the untreated group (P<0.05). On the seventh day of wound healing, Collagen I of the rats in both Shengji Huayu Recipe-treated group and Shengji Recipe-treated group was higher than that in the untreated group (P<0.05), and collagen III of the rats in both Shengji Recipe-treated group and Huayu Recipe-treated group was higher than that in the untreated group (P<0.05). CONCLUSION: Resolving stagnation and promoting granulation therapy can promote the wound healing in rats.
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<p><b>OBJECTIVE</b>To establish an effective laboratory examination system for carrier detection and prenatal diagnosis of haemophilia Alpha(HA) with a variety of molecular biological methods which are simple,rapid and easy to use.</p><p><b>METHODS</b>Detection of inversion involving intron 22 in the FVIII gene was completed by long distance polymerase chain reaction (PCR) and linkage analysis was performed by using several genetic polymorphisms including an intragenic Bcl I RFLP, 2 STRs and an extragenic St14 VNTR.</p><p><b>RESULTS</b>Intron 22 inversion was observed in 10 out of the 21 (47.6%) pedigrees examined. Prenatal diagnosis was completed in 3 pedigrees. A further combination of the four intragenic and extragenic polymorphic loci gave an informative rate of 94.7%.</p><p><b>CONCLUSIONS</b>Female relatives in HA families with inversion can be detected with direct diagnostic procedure. The application of long distance PCR makes the detection much more simple and rapid. For families without inversions,it is easier and more cost-effective to undertake linkage analysis of genetic polymorphism based on PCR.</p>
Subject(s)
Female , Humans , Pregnancy , Chromosome Inversion , Genetic Carrier Screening , Hemophilia A , Diagnosis , Genetics , Introns , Minisatellite Repeats , Polymorphism, Restriction Fragment Length , Prenatal DiagnosisABSTRACT
<p><b>OBJECTIVE</b>To explore gene defect of hereditary coagulation factor XIII deficiency.</p><p><b>METHODS</b>PCR and gene sequencing or ARMS-PCR were used to detect the FXIIIA gene of peripheral white blood cell (PBC) from two Chinese hereditary coagulation factor XIII deficiency family members and 60 normal subjects respectively. The level of FXIIIA gene mRNA was tested by RT-PCR.</p><p><b>RESULTS</b>(1) Nucleotide sequence analysis of the two probands' and their family members' DNA revealed that all of the three patients had homozygous missense mutation in FXIII A subunit gene. Proband 1 had a C to G transition at nucleotide (nt) 1 241 in exon 10 and proband 2 and his sister a C to T transition at nt 232 in exon 3 of FXIII A gene, which resulted in the substitution of Ser413 with Trp and Arg 77 with Cys, respectively. Family study showed that the two mutations were inherited from the parents who were correspondingly heterozygotes at nt 1 241 or nt 232. (2) The two mutations were not found in the normal subjects. (3) The FXIIIA gene mRNA level in the two probands was a little decreasing.</p><p><b>CONCLUSION</b>It is the two novel mutations that results in FXIIIA deficiency. The two mutations of FXIIIA gene may affect its function or alter protein folding. The defective FXIII which is unstable and degraded rapidly in cytoplasm may be the main cause of FXIII deficiency.</p>
Subject(s)
Child , Female , Humans , Blood Coagulation Disorders, Inherited , Genetics , Exons , Genetics , Factor XIII , Genetics , Factor XIII Deficiency , Genetics , Heterozygote , Pedigree , Point Mutation , Polymerase Chain Reaction , MethodsABSTRACT
<p><b>OBJECTIVE</b>To identify the mutation of coagulation factor VII (F VII) gene in two pedigrees with hereditary F VII deficiency.</p><p><b>METHODS</b>F VII gene mutations were analysed in two propositi and their family members by direct DNA sequencing. Allele specific PCR and PCR combined with restricted enzyme digestion were used to confirm the detected mutations.</p><p><b>RESULTS</b>Two gene mutations were detected in the propositus of pedigree A: G to C transition at position 6390 resulting in Trp40Cys and G to A at 11496 resulting in Arg353Gln, both are heterozygotes. The heterozygosity for polymorphism Arg353Gln was confirmed with the restriction enzyme Msp I digestion in his mother. In the propositus of pedigree B, there was a T to G transition at position 11482 resulting in His348Gln, heterozygosity of which was confirmed with Nsp I digestion in the propositus and his daughter. G to T transition at position 11514 resulting in Thr359Met was also found in the propositus of pedigree B, and the heterozygosity for Thr359Met was confirmed with allele specific PCR in the propositus and his son.</p><p><b>CONCLUSION</b>Three missense mutations were found in two pedigrees with hereditary F VII deficiency. A novel Trp40Cys mutation was reported for the first time.</p>
Subject(s)
Female , Humans , Male , Factor VII , Genetics , Factor VII Deficiency , Genetics , Heterozygote , Mutation, Missense , Pedigree , Point Mutation , Polymerase Chain Reaction , MethodsABSTRACT
<p><b>OBJECTIVE</b>To accomplish a kind of therapeutic gene for hemophilia A, and observe the expression of human factor VIII (hF VIII) in vivo.</p><p><b>METHODS</b>Human clotting factor VIII cDNA with B-domain deleted (Delta760aa approximately 1639aa) was inserted into vector pRC/RSV to form pRC/RSV-hF VIII BD, which conjugated with in vivo liposome transfection reagent (DOTAP-Cholesterol) to accomplish a kind of therapeutic gene, pRC/RSV-hF VIII BD-DOTAP-Cholesterol. Mice were injected with pRC/RSV-hF VIII BD-DOTAP-Cholesterol i.m. and sacrificed 48 hours, 10 days, 20 days, 30 days, 40 days and 50 days later, respectively. Tissues such as heart, liver, spleen, lung, kidney and muscle were harvested, the distribution and transcription as well as expression of hF VIII BD cDNA were detected by means of PCR, RT-PCR and immunohistochemistry techniques. In addition, the antigen and antibody of hF VIII in plasma were measured.</p><p><b>RESULTS</b>There was high expression of hF VIII in plasma and tissues at the 48(th) hour after injection. On day 10, antigen level of hF VIII in plasma reached its peak, 17.55 ng/ml, and gradually reduced later. The antibody of hF VIII in plasma emerged on day 10 after injection, and increased and gradually reached 37.06 U/ml on day 50 after injection. PCR, RT-PCR and immunohistochemistry showed that hF VIII BD cDNA and its transcription as well as expression existed in all kinds of tissues, and lasted longer in spleen, lungs and kidneys than in heart, liver and muscle.</p><p><b>CONCLUSION</b>Therapeutic gene, pRC/RSV-hF VIII BD-DOTAP-Cholesterol, produced by combination of pRC/RSV-hF VIII BD and DOTAP-Cholesterol liposome can express human F VIII successfully in vivo, which lays an experimental foundation for curing hemophilia A by gene-drug in clinic.</p>
Subject(s)
Animals , Humans , Mice , DNA, Complementary , Disease Models, Animal , Factor VIII , Genetics , Therapeutic Uses , Gene Expression , Genetic Therapy , Genetic Vectors , Hemophilia A , Therapeutics , Liposomes , Mice, Inbred BALB C , Tissue Distribution , TransfectionABSTRACT
<p><b>OBJECTIVE</b>To investigate the plasma levels of coagulation factor VII (FVII) and polymorphisms of FVII gene in patients with coronary heart disease (CHD), and evaluate the effect of plasma FVII levels and FVII gene polymorphisms on CHD.</p><p><b>METHODS</b>Plasma FVIIa, FVII: Ag and FVIIc were measured and polymorphisms of FVII gene were analyzed in 149 control cases and 60 CHD cases, including 33 acute myocardial infarction (AMI) cases by a combination of polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and agarose gel electrophoresis.</p><p><b>RESULTS</b>FVIIa, FVIIc in AMI group were significantly higher than that in control group, but FVII: Ag wasn't. There were no significant difference in plasma FVIIa, FVII: Ag and FVIIc between CHD and control group. The IVS7 genotypic frequency in AMI group was significantly different from that in control group. There was no significant difference in genotypic frequencies and allelic frequencies in other polymphism sites. FVII: Ag was significantly higher in -402A homozygote than that in -402G homozygote.</p><p><b>CONCLUSIONS</b>Increased FVII levels, especially FVIIa and FVIIc in plasma, may contribute to coronary artery thrombosis. There was significant difference in IVS7 genotype frequency between control and AMI groups, but the rest weren't significantly different. FVII: Ag was significantly higher in -402A homozygote than that in -402G homozygote. Polymorphism of -402 G/A may play an indirect role in AMI by regulating plasma FVII levels.</p>
Subject(s)
Female , Humans , Male , Middle Aged , Coronary Disease , Blood , Genetics , Factor VII , Genetics , Genotype , Polymerase Chain Reaction , Polymorphism, Genetic , Polymorphism, Restriction Fragment LengthABSTRACT
<p><b>OBJECTIVE</b>To explore the effect and mechanism of sodium butyrate on expression of human clotting factor VIII in vitro.</p><p><b>METHODS</b>Mouse NIH/3T3 cell line was transfected with recombinant plasmid vector pRC/RSV-BDD-hFVIII, which enclosed B-domain deleted (760aa approximately 1 639aa) human factor VIII cDNA (BDD-hFVIII cDNA). Then cells were incubated in Dulbecco's modification of Eagle's medium (DMEM) containing sodium butyrate for 24 hours, hFVIII: C and hFVIII: Ag in the cell culture medium were measured by ELISA assay and one-stage method, respectively. In addition, the effect of sodium butyrate on transcription of cDNA encoding the whole hFVIII, heavy and light chain of hFVIII was also investigated by means of run-on assay.</p><p><b>RESULTS</b>After stimulation of sodium butyrate, the levels of hFVIII: C and hFVIII: Ag increased 70% than those of control. Run-on assay showed that sodium butyrate enhanced the transcription of cDNA which encoded heavy chain of hFVIII.</p><p><b>CONCLUSION</b>Sodium butyrate can improve the expression of hFVIII through enhancing the transcription of hFVIII heavy chain encoding cDNA. It demonstrated that sodium butyrate had potential utility in inducing the expression of hFVIII in vitro.</p>
Subject(s)
Animals , Humans , Mice , 3T3 Cells , Butyrates , Pharmacology , Factor VIII , Genetics , Gene Expression RegulationABSTRACT
<p><b>OBJECTIVE</b>To test whether splicing overlapping extension(SOE) method can be a tool for obtaining rare fusion gene's transcripts and to study the tumorigenic capacity of a novel fusion gene AML1-MTG16.</p><p><b>METHODS</b>SOE method was used to obtain AML1- MTG16 fusion gene's transcripts. MTG16, AML1-MTG16 and AML1-MTG16 without III,VI conserved domains of MTG16 segment were inserted into pEGFP- C1,pDsRed-N1 vector respectively,then transfected NIH3T3 cell line by lipofection. Forty-eight hours later, the transfected cells were examined by laser-scanning confocal microscopy. Stable transfected cells were obtained by G418 500ug/ul selection for one month. Growth curve, soft agar colonies formation tumorigenesis in nude mice were done to compare the difference between stable transfected cells.</p><p><b>RESULTS</b>Recombined AML1-MTG16 by SOE contained its CDS. NIH3T3 expressing AML1-MTG16 had a faster proliferation in medium, colony growth in soft agar. AML1-MTG16 expression cells also induced tumors formation following injection into nude mouse. MTG16,AML1-MTG16 and AML1-MTG16 without III,VI conserved domains of MTG16 were colocalized in the nucleus of cotransfected NIH3T3 cells under the examination of laser-scanning confocal microscope.</p><p><b>CONCLUSION</b>SOE is an effective method to get rare fusion gene's transcripts. AML1-MTG16 plays an important role in leukemogenesis. MTG16 may also have a carcinogenic property within the AML1-MTG16 fusion gene. Carcinogenic property of AML1-MTG16 is restricted to its localization in the nuclear matrix. N terminal of MTG16 may play an important part in the carcinogenic activity of AML1-MTG16.</p>
Subject(s)
Animals , Mice , 3T3 Cells , Transplantation , Cell Division , Genetics , Cell Transformation, Neoplastic , Genetics , Cell Transplantation , Core Binding Factor Alpha 2 Subunit , Green Fluorescent Proteins , Luminescent Proteins , Genetics , Metabolism , Mice, Nude , Microscopy, Confocal , Neoplasms, Experimental , Genetics , Pathology , Oncogene Proteins, Fusion , Genetics , Plasmids , Genetics , Recombinant Fusion Proteins , Genetics , Metabolism , Time Factors , Transcription Factors , Genetics , TransfectionABSTRACT
Apoptosis, an intrinsic cell death model which is regulated by organisms, is essential for the maintenance of tissue homeostasis in multicellular organisms. Recently, researchers on cell apoptosis have paid more attention on mitochondrion than on nucleus. Different death stimulus induces opening of PT pore, degradation of mitochondrial transmembrane potential, activation of caspase and inducing cell apoptosis. Bcl-2 and Bcl-XL inhibit cell apoptosis via mitochondrion while Bax, Bak and Bad induce apoptosis by means of regulation of mitochondrion.