Objective To report the clinical and genetic characteristics of a dentatorubralpallidoluysian atrophy (DRPLA) pedigree with an onset of cognitive impairment.Methods Clinical data of this pedigree was collected.The numbers of CAG repeats in the exon 5 of atrophin-1 (ATN1) gene were analysed in the proband and the other 4 healthy family individuals.The polymerase chain reaction (PCR) products of the proband underwent cloning-sequencing using an original TA cloning kit.Results There were 5 patients in this family,4 with onset in adult and one in childhood.The proband had an onset manifestation of cognitive impairment,while the other 3 adult patients presented with ataxia.The two-year-old child in the pedigree had myoclonic epilepsy.The proband had 61 CAG repeats in the exon 5 of ATN1 gene.After TA cloning-sequencing of the proband ' s PCR products,there were 2 different numbers of CAG repeats,including 61 and 64.Conclusions The clinical manifestations of DRPLA can have obvious heterogeneity in one family.Some patients present with cognitive impairment.It is very important to test the numbers of CAG repeats of ATN1 gen for DRPLA diagnosis.Somatic mosaicism may be also observed in Chinese DRPLA patients.

AIM:To analyze the effects of oxymatrine(OMT) on the quantity of murine regulatory T cells(Tr cells) in the peripheral blood and mouse lymphocyte proliferation stimulated by Con A,and to probe into the immunological mechanism that OMT treats allergic contact dermatitis(ACD).METHODS:An ACD mouse model stimulated by dinitrofluorobenzene(DNFB) was established.Different dosages of OMT,PBS and hydrocortisone(HCT) were intraperitoneally injected(IP) into the mice.Blood samples were collected at 1 d,7 d,14 d,21 d and 28 d,then the T cells were isolated and marked with anti-CD3,anti-CD4,anti-CD25 three-colored immune fluorescence antibody to detect the quantity of CD4+CD25+ T cells with flow cytometry.The fluorescence intensity changes of lymphocytes which were isolated from mouse's lymph node and co-stimulated by polyclonal stimulator Con A and OMT were examined by carboxyfluorescein diacetate succinimidyl ester(CFDA-SE) staining and flow cytometry.RESULTS:OMT at concentrations of 500,125 and 31 mg/L had the ability to restrain the proliferation of lymphocytes from lymph node in a dose dependent manner.However,OMT at concentrations of 16,8,4 and 2 mg/L promoted the proliferation of T lymphocytes from lymph node,but was not obviously dependent on its concentration.Intraperitoneal injection of OMT increased the numbers of CD4+CD25+T cell in peripheral blood obviously(P

AIM: To explore the inhibitory effect of oxymatrine (OMT) on the allergic contact dermatitis (ACD) stimulated by dinitrofluorobenzene (DNFB) and its effects on the proliferation of the lymphocytes. METHODS: ① An ACD mouse model was established by stimulation with DNFB, and then the mice were injected intraperitoneally with different dosages of OMT, PBS and hydrocortisone (HCT) respectively, the swelling degree of their auricles was examined. ② Carboxyfluorescein diacetate, succinimidyl ester (CFDA-SE) dye and flow cytometer were used to examine the fluorescence intensity changes of lymphocytes stimulated by polyclonal stimulator ConA and OMT. RESULTS: ① compared with PBS group, OMT possessed the strong inhibitory effect on the ACD caused by DNFB in a dose-dependent manner, and its inhibitory effect was equivalent to the HCT of the same dosage with fewer side effects. ② In vitro experiments proved that OMT (500, 125 and 31 mg/L) had the ability to restrain the proliferation of lymphocytes of mouse. CONCLUSION: OMT possesses an inhibitory effect on the ACD induced by DNFB, and OMT is a kind of immunosuppressor.

Objective To report the clinical,myopathological and genetic features of a patient with distal myopathy caused by caveolin-3 (CAV3) deficiency.Methods The patient was a 27-year-old female.She had an onset symptom of asymmetric lower extremities weakness.The proximal limb-girdle muscles were involved subsequently.Clinical data of this patient were collected.The leg muscle magnetic resonance imaging (MRI) and an open biopsy of left tibialis anterior muscle were performed.In addition to histological,enzyme histochemical staining and ultrastructural examination,immunohistochemical staining with antibody against CAV3 was done.CAV3 gene was analyzed in the patient and her parents.Results Tl-weighted enhanced skeletal muscle MRI of the lower limbs showed the abnormal signal in distal and proximal muscles.Muscle biopsy showed moderate dystrophic changes and immunostaining for CAV3 showed reduced plasmalemma in the muscle fibers.Gene analysis disclosed a heterozygous c.136G ＞ A (p.Ala46Thr)mutation in the CAV3 gene,and the patient's parents did not have this mutation.Conclusions We report a distal myopathy case caused by c.136G ＞ A (p.Ala46Thr) mutation in the CAV3 gene,who had an onset symptom of asymmetric lower extremities weakness.The proximal limb-girdal muscles were also involved.This would help clinical doctors to know more about this rare myopathy.

OBJECTIVE: To evaluate enantiomeric separation methods for beta-blocking agents and analogs. METHODS: Enantiomeric separation of racemates of 11 beta-blocking agents and their analogs was performed using chiral stationary phases and 2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl isothiocyanate (GITC). RESULTS: These beta -blocker racemates were separated into enantiomers in one or several chormatographic states such as propranolol, bisoprolol, metoprolol, celiprolol, carvedilol, sotalol, propafenone, ephedrine, and zomitriptan. Temperature had a significant effect on the resolution of the drugs when using chiralcel OD. Lower temperatures were associated with higher resolutions. CONCLUSION: When separating beta-blocking agents and their analogs, Chiralcel OD, Chiralpak AD, Chiral stationary phases and GITC chiral derivative reagents have complementary functions.

This study aimed to construct recombinant adenovirus expressing Epstein-Barr nuclear antigen 1 (EBNA1) against nasopharyngeal carcinoma (NPC). The C-terminal region fragment of the ebna1 gene of Epstein-Barr virus was amplified from the standard strain B95-8 by polymerase chain reaction (PCR). The gene fragment was inserted into the pDC316 shuttle plasmid using the EcoRI and BgIII restriction enzyme sites. The pDC316-ebna1 shuttle plasmid and pBHG helper plasmid were cotransfected into HEK293 cells after sequencing. The soluble protein was extracted from HEK293 cells, which caused apparent cytopathic effects. The transcription and expression of the ebna1 gene were confirmed using flow cytometry and Western blotting. rAd-ebna1 titers were measured by the TCID50. rAd-ebna1 was injected into BALB/c mice at a dose of 2 x 10(8) VP per mouse, EBNA1 epitope-specific responses were measured at 1st, 2nd, 4th and 8th weeks post-immunization. The target fragment of ebna1 (939 bp) was obtained by PCR, and was in consensus with the sequence from the standard strain B95-8. Cytopathic effects were observed after the pDC316-ebna1 shuttle plasmid and pBHG helper plasmid were cotransfected into HEK293 cells. rAd-ebna1 was successfully recombined in HEK293 cells. EBNA1 protein was detected in HEK293 cells, rAd-ebna1 titers reached 10(8) TCID50/mL. Specific responses to CD4+ epitopes of EBNA1 were detected in the immunized mice. In conclusion, rAd-ebna1 was successfully constructed and induced specific responses to CD4+ epitopes of EBNA1 in immunized mice.
Adenoviridae ; genetics ; immunology ; Animals ; CD4-Positive T-Lymphocytes ; immunology ; virology ; Epstein-Barr Virus Infections ; immunology ; prevention & control ; virology ; Epstein-Barr Virus Nuclear Antigens ; administration & dosage ; genetics ; immunology ; Genetic Vectors ; genetics ; immunology ; Herpesvirus 4, Human ; genetics ; immunology ; Humans ; Male ; Mice ; Mice, Inbred BALB C ; Viral Proteins ; administration & dosage ; genetics ; immunology

Aged ; Female ; Femoral Neck Fractures ; diagnostic imaging ; surgery ; Fracture Fixation, Internal ; methods ; Humans ; Male ; Middle Aged ; Radiography ; Retrospective Studies ; Treatment Outcome

Objective To explore the effects of Tongyuan acupuncture combined with herbal medicine and moxibustion on the behavior and hypothalamic-pituitary-adrenal axis-related hormone levels in postpartum de-pression rats.To explore the therapeutic mechanism of this method in the treatment of postpartum depression.Meth-ods A total of 120 SD rats were randomly divided into 6 groups(n=20).The blank group was not treated.The sham operation group was performed without surgery to remove the ovaries. The model group,drug group,routine acupuncture group and experimental group were established in the model of postpartum depression.After modeling successfully,the model group did not interfere with any treatment. The rats in treatment group were treated with fluoxetine hydrochloride. The acupuncture group was treated with acupuncture of Baihui and Taichong. The rats in the experimental group were treated with the method of acupuncture and moxibustion.At the end of the second and fourth week after the initiation of intervention,10 rats were randomly selected from each group.They were sacrificed and the levels of serum corticotropin-releasing hormone(CRH),adrenocorticotropic hormone(ACTH),cortisol (Cor)were measured.Results The levels of CRH,ACTH and Cor in the model group were higher than those in the blank group(P<0.05).The levels of serum CRH,ACTH and Cor in the drug group,routine acupuncture group and experimental group were lower than those in the model group(P<0.05).The levels of serum ACTH and Cor in the experimental group were lower than those in the drug group and routine acupuncture group(P<0.05).There was no significant difference in serum CRH,ACTH and Cor level between the experimental group and blank group(P >0.05).Conclusions Tongyuan acupuncture combined with salt moxibustion and Shenque have significant effects on the depressive symptoms and serum levels of CRH,ACTH and Cor in postpartum depression rats,which may be re-lated to the regulation of hypothalamic-pituitary-adrenal axis-related hormone levels.

OBJECTIVETo explore the growth inhibiting effects on human bladder cancer by antisense RNA targeting the proliferating cell nuclear antigen (PCNA) gene.

METHODSThe eukaryotic expression vector for antisense PCNA cDNA was constructed and transferred into a bladder cancer EJ cell line. The PCNA expression in the cancer cells was detected by RT-PCR and Western blotting assays. The in vitro proliferation activities of the transferred cells were observed by growth curve, tetrazolium bromide (MTT) colorimetry, tritiated thymidine ((3)H-TdR) incorporation, flow cytometry and clone formation testing, while its in vivo anti-tumor effects were detected on nude mice allograft models.

RESULTSAfter the antisense vector, pLAPSN, was transferred, cellular PCNA expression was inhibited at both protein and mRNA levels. The growth rates of EJ cells were reduced from 27.91% to 62.07% (P < 0.01), with an inhibition of DNA synthesis rate by 52.31% (P < 0.01). Transferred cells were blocked at G(0)/G(1) phases in cell-cycle assay, with the clone formation ability decreased by 50.81% (P < 0.01). The in vivo carcinogenic abilities of the transferred cancer cells were decreased by 54.23% (P < 0.05).

CONCLUSIONSAntisense PCNA gene transfer could inhibit the growth of bladder cancer cells in vitro and in vivo, which provided an ideal strategy for gene therapy of human cancers.

Animals ; Cell Division ; genetics ; Gene Transfer Techniques ; Genetic Therapy ; methods ; Genetic Vectors ; Humans ; Mice ; Mice, Nude ; Proliferating Cell Nuclear Antigen ; genetics ; RNA, Antisense ; genetics ; Tumor Cells, Cultured ; Urinary Bladder Neoplasms ; pathology

OBJECTIVETo develop an RP-HPLC method for assay of pilocarpine in rabbit ocular aqueous humor.

METHODSThe RP-HPLC method was performed on a column of ODS-C(18) with the mobile phase consisting of 0.5% of triethylamine (TEA) of phosphate solutions (10 mmol/L, pH 2.5) and acetonitrile (98/2,v/v). The detection wavelength was 215 nm and flow rate was 1.0 ml/min. Ninety albino rabbits were divided into 3 groups (30 in each):group 1 received 50 microl of eye drops containing 1% generic pilocarpine, group 21% mixture pilocarpine solution consisting of aqueous sample and liposome and group 31% liposome pilocarpine, respectively. The aqueous humor was withdrawn at 5, 10, 30, 40, 60, 90, 120, 180, 240 and 360 min. Pilocarpine was extracted from aqueous humor with dichloromethane.

RESULTThe linear calibration curve was obtained in the concentration range of 0.1 - 20 microg/ml. The average recovery was (68.1+/-2.7)% (n=9). Inter-day and intra-day RSD were 4.33% and 2.87%, respectively. In three formations 1% liposome pilocarpine was the best for the areas under curve and measurable amounts.

CONCLUSIONThe RP-HPLC method is simple and reliable for pilocarpine measurement in ocular aqueous. Liposome formulation can significantly increase the bioavailability of pilocarpine in ocular aqueous.

Animals ; Aqueous Humor ; chemistry ; Chemistry, Pharmaceutical ; Chromatography, High Pressure Liquid ; Pilocarpine ; administration & dosage ; analysis ; Rabbits