尿路清治疗解脲支原体感染非淋菌性尿道炎(宫颈炎)临床研究

目的:观察尿路清治疗解脲支原体(Uu)感染非淋菌性尿道炎(宫颈炎)的临床疗效。方法:将119例患者随机分为3组。A组39例以尿路清(由白花蛇舌草、土茯苓、崩大碗、黄柏、黄芪、旱莲草、地肤子等组成)治疗,B组35例以强力霉素治疗,C组45例以尿路清加强力霉素治疗。结果:总有效率A组为71.8%,B组为48.6%,C组为88.9%,A组与B组、C组与A组比较,差异均有显著性意义(P<0.05);C组与B组比较,差异有非常显著性意义(P<0.01)。治疗后Uu阴转A组20例(51.3%),B组15例(42.9%),C组33例(73.3%),A组与B组比较,差异无显著性意义(P>0.05);C组与A组比较,差异有显著性意义(P<0.05);C组与B组比较,差异有非常显著性意义(P<0.01)。结论:尿路清治疗Uu感染非淋菌性尿道炎(宫颈炎)有较好的临床疗效。

野兔的人工养殖

野兔以其肉质细嫩香醇、味美而倍受人们青睐。野兔一般饲养3个月即可上市出售,是致富的一条好路子。现将人工饲养技术介绍如下。 野兔生存的基础是野生环境。人工养殖野兔,既要模拟野生环境,又要考虑饲养管理需要。一是造窝。野兔不打洞,因此要在饲养场内堆放些麦秸杆、花生秧、玉米秸秆等,

Establishment of an artificial β-cell line expressing insulin under the control of doxycycline

AIM: Artificial beta-cell lines may offer an abundant source of cells for the treatment of type I diabetes, but insulin secretion in beta-cells is tightly regulated in physiological conditions. The Tet-On system is a &quot;gene switch&quot; system, which can induce gene expression by administration of tetracycline (Tet) derivatives such as doxcycline (Dox). Using this system, we established 293 cells to an artificial cell line secreting insulin in response to stimulation by Dox. METHODS: The mutated proinsulin cDNA was obtained from plasmid pcDNA3.1/C-mINS by the polymerase chain reaction (PCR), and was inserted downstream from the promoter on the expression vector pTRE2, to construct a recombined expression vector pTRE2mINS. The promoter on pTRE2 consists of the tetracycline-response element and the CMV minimal promoter and is thus activated by the reverse tetracycline-controlled transactivator (rtTA) when Dox is administrated. pTRE2mINS and plasmid pTK-Hyg encoding hygromycin were co-transfected in the tet293 cells, which express rtTA stably. Following hygromycin screening, the survived cells expressing insulin were selected and enriched. Dox was used to control the expression of insulin in these cells. At the levels of mRNA and protein, the regulating effect of Dox in culture medium on the expression of proinsulin gene was estimated respectively with Northern blot, RT-PCR, and radioimmunoassay. RESULTS: From the 28 hygromycin-resistant cell strains, we selected one cell strain (tet293/Ins6) secreting insulin not only automatically, but in response to stimulation by Dox. The amount on insulin secretion was dependent on the Dox dose (0,10,100,200,400,800 and 1000 microg.L(-1)), the level of insulin secreted by the cells treated with Dox (1000 microg.L(-1)) was 241.0pU.d(-1).cell(-1) , which was 25-fold that of 9.7pU.d(-1).cell(-1) without Dox treatment. Northern blot analyses and RT-PCR further confirmed that the transcription of insulin gene had already been up-regulated after exposing tet293/Ins6 cells to Dox for 15 minutes, and was also induced in a dose-dependent manner. However, the concentration of insulin in the media did not increase significantly until 5 hours following the addition of Dox. CONCLUSION: Human proinsulin gene was transfected successfully and expressed efficiently in 293 cells, and the expression was modulated by tetracycline and its derivatives, improving the accuracy, safety, and reliability of gene therapy, suggesting that conditional establishment of artificial beta-cells may be a useful approach to develop cellular therapy for diabetes mellitus.

Esophageal ulceration complicating doxycycline therapy

AIM;TO report present state of iatrogenic drug-induced esophageal injury(DIEI)induced by medications in a private clinic. METHODS:Iatrogenic drug-induced esophageal injury (DIEI)induced by medications has been more frequently reported.In a private clinic we encountered 36 cases of esophageal ulcerations complicating doxycycline therapy in a mainly younger Saudi population(median age 29 years). RESULTS:The most frequent presenting symptoms were odynophagia,retrosternal burning pain and dysphagia(94 %, 75 % and 56 %,respectively).The diagnosis was according to medical history and confirmed by endoscopy in all patients. Beside withdrawal of doxycycline,when feasible,all patients were treated with a proton-pump inhibitor(PPI)and a prokinetic.Thirty patients who reported to the clinic after treatment were improved within 1-7(median 1.7)days. CONCLUSION:Esophageal ulceration has to be suspected in younger patients with odynophagia,retrosternal burning pain and/or dysphagia during the treatment with doxycycline.

Doxycycline blocks gastric ulcer by regulating matrix metalloproteinase-2 activity and oxidative stress

AIM: To examine the effect of doxycycline on the activity of matrix metalloproteinases (MMPs) and oxidative stress in gastric tissues of rats following gastric injury.METHODS: Gastric ulcers were generated in rats by administration of 70% ethanol,and activity of doxycycline was tested by administration 30 min prior to ethanol.Similarly,the effect of doxycycline was tested in an indomethacin-induced gastric ulcer model.The activities and expression of MMPs were examined by zymography and Western blot analysis.RESULTS: Gastric injury in rats as judged by elevated ulcer indices following exposure to ulcerogen,either indomethacin or ethanol,was reversed significantly by doxycycline.Indomethacin-induced ulcerated gastric tissues exhibited about 12-fold higher proMMP-9 activity and about 5-fold higher proMMP-3 activity as compared to control tissues.Similarly,ethanol induced about 22-fold and about 6-fold higher proMMP-9 and proMMP-3 activities,respectively,in rat gastric tissues.Both proMMP-9 and MMP-3 activities were markedly decreased by doxycycline in ulcerogen treated rat gastric tissues.In contrast,the reduced MMP-2 activity in ulcerated tissues was increased by doxycycline during ulcer prevention.On the other hand,doxycycline inhibited significantly proMMP-9,-2 and -3 activities in vitro.In addition,doxycycline reduced oxidative load in gastric tissues and scavenged H2O2 in vitro.Our results suggest a novel regulatory role of doxycycline on MMP-2 activity in addition to inhibitory action on MMP-9 and MMP-3 during prevention of gastric ulcers.CONCLUSION: This is the first demonstration of dual action of doxycycline,that is,regulation of MMP activity and reduction of oxidative stress in arresting gastric injury.

The Inhibitory Effect of Endostatin and Doxycycline Administration on B16 Melanoma Angiogenesis and Cellular Proliferation

OBJECTIVE To investigate the effect of endostatin and doxycycline on melanoma cellular proliferation and tumor angiogenesis.METHODS The effects of endostatin and doxycycline were studied in mice transplanted with B16 melanoma cells. The mice were divided into 4 groups that were treated as follows： endostatin treatment （E group）, doxycycline treatment （D group）, endostatin plus doxycycline trearment （DE group）, controls （C group） received no treatment. Following 9 days of treatment the tumor tissue was removed to compare the differences in the tumor necrotic rate and micro-vessel density （MVD） among the different groups. Immunohistochemical staining was conducted to detect the expression of proliferating cell nuclear antigen （PCNA） in the different groups.RESULTS The MVD of the 3 experimental groups was significantly less than the control group, （F = 10.888, P 〈 0.05）, indicating that doxycycline and endostatin can inhibit tumor angiogenesis by decreasing the tumor blood supply. This effect results in inhibition of tumor cellular proliferation and promotion of tumor cell necrosis. The tumor cell necrotic rate of the 3 experimental groups were all significantly higher than the C group （F = 7.229, P 〈 0.05） and the difference between the DE and C groups also was statistically significant. PCNA expression in all 3 experimental groups was statistically less than the C group （F = 17.729, P 〈 0.05）.CONCLUSION The combined use of endostatin and doxycycline in vivo can influence PCNA expression and angiogenesis in melanoma, and significantly inhibit melanoma cellular proliferation.

A SINGLE TETRACYCLINE-REGULATED VECTOR DEVISED FOR CONTROLLED INSULIN GENE EXPRESSION

Objective To construct a single plasmid vector mediating doxycycline-inducible recombined human insulin gene expr-ession in myotube cell line. Methods An expression cassette of rtTAnls driven by promoter of human cytomegalovirus and a furin-cuttable recom-bined human insulin expression cassette driven by a reverse poly-tetO DNA motif were cloned into a single plasmid vector (prTR-tetO-mINS). The prTR-tetO-mINS and pLNCX were co-transfected into a myotube cell line (C2C12) and pLNCX vector were used as a control. After selection with G418, the transfected cells were induced with doxycycline at concentra-tions of 0, 2, and 10 μg/mL. RT-PCR was used to determine expression levels of recombinant insulin mRNA at the 5th day. Insulin production in cell cultures medium (at different incubation time) and cell extracts (at the 7th day) were analyzed with human pro/insulin RIA kits. Results Immune reactive insulin (IRI) level in cell medium was found increased at 24 hours of doxycycline incubation, and still increased at the 5th day. After withdrawn of doxycycline, IRI decreased sharply and was at baseline three days later. IRI and human insulin mRNA levels were positively related to different levels of doxycycline. A 25-fold increase in IRI was found against background expression at the 7th day. Conclusion Human insulin expression can be successfully regulated by doxycycline and the background was very low. This single tet-on insulin expression system may provide a new approach to a controlled insulin gene therapy in skeletal muscle.

Construction and identification of immortalized rat astrocyte cell line expressing enkephalin

Objective:To provide a sound cell source for further ex-vivo gene therapy for chronic pain,we attempt to develop an immortalized rat astrocyte cell line that expresses enkephalin regulated by doxycycline. Methods:Retrovirus infection method was employed to develop an immortalized rat astrocyte cell line that could express enkephalin regulated by doxycycline.The hPPE gene expression level of immoralized astroyte cells(IAC)/ hPPE was detected by RT-PCR,indirect immunofluorescence staining and radioimmunoassay. Results:IAC carrying Tet-on system transfected with preproenkephalin gene could secrete enkephalin that was regulated by doxycycline in a dose-dependent manner and hPPE gene activation could be repeated in on-off-on cycles through administration or removal of doxycycline. Conclusion:An immortalized rat astrocyte cell line that secrete enkephalin under the control of doxycycline is established successfully,which provides a research basis for transgenic cell transplantation for analgesia.