商陆抗病毒蛋白对感染细胞模型中HCV复制的影响

目的探讨商陆抗病毒蛋白(PAP)对丙型肝炎病毒(HCV)感染细胞模型中 HCV 的抑制作用。方法用 HCV 阳性血清感染人肝癌细胞株 HepG2细胞.制成 HCV感染细胞模型,用不同浓度的 PAP 干预该模型;利用荧光定量 PCR 法分别于药物干预后48h、96h、144h 检测培养细胞及上清液中 HCV RNA 含量,同时以不同浓度干扰素(IFN)处理该模型作为对照。结果 PAP 处理 HepG2感染 HCV细胞模型后,细胞内 HCV RNA 含量在48h、96h、144h 各组间与对照组相比有显著差异(P<0.01)。在第48h,HCVRNA 含量100μg/ml 组和10μg/ml 组<1μg/ml 组<0.1μg/ml 组<0.01μg/ml 组及对照组;在第96h,HCV RNA含量100μg/ml 组和10μg/ml 组<1μg/ml 组<0.1μg/ml 组和0.01μg/ml 组<对照组;在第144h,HCVRNA含量100μg/ml 组及10μg/ml 组<1μml 组<0.1μg/ml 组、0.01μg/ml组及对照组。培养上清液中 HCV RNA 含量各组间在48h时与对照组相比差异无显著性(P>0.05),第96h,培养上清液中 HCV RNA 含量各组间差异显著(P<0.05)。100μg/ml组及10μg/ml 组<1μg/ml 组<0.1μg/ml 组、0.01μg/ml 组及对照组;第144h 时,培养上清液中 HCV RNA 含量各组间差异显著(P<0.01),100μg/ml 组、10μg/ml 组及1μg/ml 组<0.1μg/ml 组、0.01μg/ml 组及对照组。PAP 对 HCV 的抑制作用随 PAP 浓度的增加而增强,以100μg/ml 组对 HCV的抑制作用最强。IFN 干预该模型亦得出相似结果,以3.0×10~4IU/ml 组对 HCV 抑制作用最强。PAP 与 IFN 均以第48h 的抑制效果最好,在48h、96h 及144h时,PAP 对 HCV的抑制作用显著高于 IFN(P<0.01)。实验浓度的 PAP 未引起细胞死亡或脱壁,对细胞的形态、生长也无明显影响。结论 PAP 对 HCV 复制有明显的抑制作用,且作用强于IFN。实验浓度的 PAP 对细胞形态及生长无明显影响。     

商陆抗病毒蛋白抗HCV作用的初步研究

目的　探讨商陆抗病毒蛋白 (PAP)对HCV的抑制作用。方法　用不同浓度PAP处理HCV感染细胞模型 ,用荧光定量PCR检测HCVRNA。结果　HepG2 细胞内HCV持续表达达 3 0d以上。PAP处理HCV HepG2 细胞后 ,各组细胞内HCVRNA含量在 48h、96h和 14 4h差异非常显著 (P <0 .0 1)。培养上清液HCVRNA含量在 48h时 ,各组间无明显差异 (P >0 .0 5 ) ,但在 96h和 14 4h时 ,各组间差异非常显著 (P <0 .0 1)。此种抑制作用随PAP浓度的增加而增强。结论　PAP具有抑制HCV复制和表达的作用。     

慢性丙肝患者血清干扰素抗体检测及其临床意义

选择94例慢性丙型病毒性肝炎(慢性丙肝)患者，采用α2b—干扰素(α2b—IFN)治疗，动态检测其血清抗—HCV、HCV—RNA及α2b—IFN—IgG表达水平。结果显示α2b—IFN治疗后抗—HCV、HCV—RNA阴转率分别为32．98％(31／94)和38．30％(36／94)，常规治疗组治疗后抗—HCV、HCV—RNA阴转率分别为10．OO％(4／40)和15．OO％(6／40)，两者相比差异有显著性(P＜O．01)。α2b—IFN治疗后抗—IFN—IgG阳检率为5．32％(5／94)，与常规治疗组相比差异无显著性(P＞O．05)。认为α2b—IFN对抗—HCV、HCV—RNA阴转具有肯定疗效，且优于常规治疗，是治疗慢性丙肝的有效药物之一。α2b—IFN虽有一定的免疫原性，但在治疗过程中仅诱导机体产生低水平的抗α2b—IFN—IgG，对IFN—α2b的抑制作用较弱。     

慢性丙型肝炎干扰素治疗后复发患者干扰素联合利巴韦林再治疗

目的 探讨干扰素（IFN）治疗后复发的慢性丙型肝炎（CHC）患者对IFN联合利巴韦林再治疗的应答情况及影响因素。方法 100例IFN治疗后复发的CHC患者中，50例使用聚乙二醇干扰素α-2a（PEG—IFNα-2a），50例使用重组人干扰素α-1b（CIFNα—1b），均联合利巴韦林再治疗，联合治疗48周，停药随访24周，分析HCVRNA载量、病毒基因型、药物种类对联合治疗疗效的影响。结果 100例复发患者联合再治疗后，36．00％取得持续病毒学应答（SVR），其中PEG-IFNα-2a组48．00％取得SVR，显著高于CIFNα—1b组（24．00％，P〈0．05）。56例低病毒载量（HCV-RNA〈1×10^5拷贝／mL）患者中，PEG—IFNα-2a组28例，其中57．14％取得SVR，显著高于CIFNα—1b组（25．00％，P〈0．05）。HCV非基因1（2a或2b）型组29例，其中55．17％取得SVR，显著高于基因1型组（28．20％，P〈0．05）；在CIFNα—1b治疗组，病毒非基因1型17例患者，其中47．06％取得SVR，明显高于基因1型患者（12，12％，P〈0．01）；在基因1型组，PEG—IFNα-2a组38例，其中42．11％取得SVR，显著高于CIFNα—1b组（12．12％，P〈0.01）。结论 IFN治疗后复发的CHC患者IFN联合利巴韦林再治疗存在部分患者无应答；对于HCV病毒载量低、基因1型的复发患者，聚乙二醇干扰素联合利巴韦林再治疗疗效明显优于普通干扰素的联合治疗。     

干扰素γ对丙型肝炎病毒复制子的抑制作用

目的研究干扰素（IFN）γ直接抑制丙型肝炎病毒（HCV）复制子的作用及其与IFN α联合用药的疗效，并对可能介导IFN γ抗HCV作用的干扰素刺激基因（ISG）的表达水平进行探讨。方法建立HCV复制子细胞模型，用IFN γ或IFN γ联合IFN α处理HCV复制子细胞，通过半定量逆转录聚合酶链反应（RT-PCR）及实时定量PCR检测细胞中HCV RNA水平；western blot检测细胞中HCV非结构蛋白5A（NS5A）。结果IFN γ对HCV RNA的复制及NS5A的表达有明显抑制作用，10U／ml IFN γ可使HCV RNA较对照组减少36％，25U／ml减少80％；IFN γ对HCV RNA及NS5A蛋白的抑制作用与其剂量及作用时间呈正相关。用IFN γ预处理的细胞对IFN α抗HCV的作用更敏感，且细胞内干扰素调节因子-1（IRF-1）、干扰素刺激基因因子3 γ（ISGF3 γ）和2’5’-寡腺苷酸合成酶（2’5’-OAS）呈明显的诱导性表达。结论IFN γ能有效抑制HCV RNA的复制，并与IFN α有协同抗HCV的作用。IRF-1、ISGF3 γ和2’5’-OAS可能参与介导IFN γ抗HCV的作用。     

干扰素α联合利巴韦林治疗丙型肝炎病毒与人类免疫缺陷病毒混合感染

目的观察干扰素α联合利巴韦林对中国人丙型肝炎病毒（HCV）与人类免疫缺陷病毒（HIV）混合感染（HCV／HIV）患者的治疗效果及不良反应，并与其对单独HCV患者的治疗进行对比。方法10例HCV／HIV患者和17例HCV患者，以肌肉注射干扰素α5MU，隔日1次，口服利巴韦林0．3g，3次／d，动态观察两组的HCV RNA水平和HIV RNA水平，CD^4＋和CD8^＋ T淋巴细胞计数，主要的肝功能和血细胞指标，以及主要的不良反应。结果HCV／HIV患者与HCV患者治疗l2周和24周，HCV RNA较用药前平均分别下降1．14log（t-3．843，P〈0．01）和2．08log（f=6．564，P〈0．01），1．48log（f=6．438，P〈0．01）和2．33log（t=7．343，P〈0．01）;同时HIV RNA也较用药前平均下降了1．22log（t=3．662，P〈0．01）和1．73log（t=6．119，P〈0．01），但两组用药前后的CD4^＋、CD8^＋T淋巴细胞计数变化不大；27例患者的氨基转移酶除2例稍高以外其余均降至正常值。用药过程中部分病例发生流感样症状和消化道反应（多见于用药早期），白细胞总数和血红蛋白各有4例轻度下降；未发现明显精神神经异常和自身免疫疾病表现。结论以干扰素α联合利巴韦林治疗中国人HCV／HIV患者，用药24周的抗HCV效果不低于两种药物联合治疗单独HCV患者的效果，并有一定的抗HIV作用。两组的生化反应和不良反应差异无统计学意义。     

干扰素α抗病毒活性的实验研究

目的比较不同亚型干扰素α（IFNα2b、IFNα2a、IFNαIb）对JAK-信号转导活化转录因子（STAT）通道中重要信号传导分子STAT1、STAT2、干扰素α受体（IFNAR）、蛋白激酶（PKR）、核糖核酸酶L（RNaseL）表达的影响，研究其抗病毒活性的差别，并评价IFNα抗病毒活性的关键性信号传导分子。方法1000U／ml的不同亚型IFNα分别作用于HepG2细胞后，用逆转录聚合酶链反应（RTPCR）方法检测HepG2细胞内STAT1、STAT2、IFNAR、PKR、RNase L mRNA表达水平。用western blot方法检测细胞内STAT1及IFNAR蛋白表达水平。结果RT-PCR的实验结果：（1）IFNα1b处理组IFNAR、STAT1、STAT2 mRNA的表达水平较IFNα2b组高，两组比较差异无统计学意义。IFNα1b、IFNα2b处理组IFNAR、STAT1、STAT2 mRNA水平明显较IFNα2a组高，差异均有统计学意义，，值分别为5．26、15．6、4．67，P值均〈0．05。（2）IFNα1b、IFNα2b、IFNα2a处理后的PKR表达水平相似，3组比较差异无统计学意义。（3）RNaseL表达量极少，无法比较不同处理组间RNase L mRNA表达水平的差异性。Western blot实验结果：（1）IFNα1b处理组IFNAR、STAT1蛋白水平较IFNα2b处理组高，两者比较差异无统计学意义，IFNα1b、IFNα2b处理组IFNAR、STAT1蛋白水平较IFNα2a处理组明显升高，差异有统计学意义，F值分别为17．7、20．1，F值均〈0．05。结论IFNα2b、IFNα2a、IFNα1b均可促进JAK—STAT信号通道中STAT1、STAT2、IFNAR mRNA及蛋白表达，IFNα1b和IFNα2b作用较强，IFNα2a作用较弱。初步表明，IFNα1b、IFNα2b的抗病毒活性较IFNα2a强，IFNAR、STAT1、STAT2可作为评价IFNα抗病毒活性的关键性指标，而PKR、RNaseL能否作为评价指标有待进一步的实验证实。     

重组干扰素α-2a,α-2b治疗慢性乙型肝炎疗效比较

探讨不同亚型α干扰素（IFNα-2α，IFNα-2b）治疗慢性乙型肝炎的疗效差异。110例慢性乙型肝炎（CHB）患者被随机分为IFNα-2α，治疗组（52例）和IFNα-2b治疗组（58例）。IFNα-2α治疗组采用IFNα-2a（因特芬）每日3MU肌注，IFNα-2b治疗组采用IFNα-2b（隆化诺）每日3MU肌注，30天后改隔日3MU肌注，疗程6月。观察两组用药3月，6月后血清ALT昨常率，HBeAg阴转率，HBV－DNA阴转率和治疗反应率。ALT复常率两组无明显差异（P＞0．05）。HBeAg阴转率：IFNα－2b治疗组（51.7%,67.2%)，明显优于IFNα－2a治疗组（32.7%,44.2%)，两组差异明显（P＜0．05）。HBV DNA阴转率：6月后IFNα－2b治疗组（58．6％），明显优于IFNα-2α治疗组（34．6％），两组差异明显（P＜0．05）。治疗后完全反应率：IFNα－2b治疗组（46．6％） ，明显优于IFNα-2α治疗组（21．2％），两组间有极显著差异（P＜0．01）。干扰素α-2b亚型治疗慢性乙型肝炎疗效明显优于干扰素α-2α，亚型。     

细胞培养实验中条件培养基相关问题探讨

目的“通过LX-2人肝星状细胞条件培养基（CM）对HepG2人肝癌细胞增殖活性的影响,探讨CM各影响因素在离体实验中的作用。方法：收集LX-2细胞第24小时、48小时、72小时条件培养基,设100％、50％、25％共3个浓度,并设1％DMEM组为对照,培养24小时、48小时、72小时;并将收集的第24小时条件培养基分装分别保存至4℃1周、-20℃1周、-80℃3个月、-20℃3个月,采用MTT法检测各组条件培养基对HepG2细胞增殖活性的影响。结果：①收集时间：第24小时、第48小时LX-2CM具有明显的促增殖作用（P〈0．01）;培养24小时后,各收集时间点的LX-2CM的促增殖作用为：第24小时〉第48小时〉第72小时（P〈0．01）;培养48小时后,第48小时〉第72小时（P〈0．01）。②添加浓度：培养24小时后,第24小时CM100％浓度、第24小时CM50％浓度、第24小时CM25％浓度、第48小时CM100％浓度均有明显的促增殖作用（P〈0．01）;第24小时CM各浓度比较,100％优于50％（P〈0．05）;第48小时CM各浓度,100％优于50％（P〈0．05）、25％（P〈0．01）。③多元线性回归分析,条件培养基收集时间与HepG2细胞增殖呈负相关,浓度与其呈正相关。④保存条件：-20℃1周CM优于-80℃3个月CM、-20℃3个月CM（P〈0．01）;-800C3个月CM优于-20℃3个月CM（P〈0．01）。结论：LX-2CM对HepG2细胞具有明显的促进增殖活性作用,其作用随着收集时间点延长、浓度减小,增殖效果逐渐降低;CM的保存条件对其作用效果有影响。     

宿主细胞因子及黏病毒抵抗蛋白A基因多态性与干扰素治疗慢性乙型肝炎疗效的相关性

目的：探讨IFN治疗慢性乙型肝炎（CHB）患者的疗效与TNF—α启动子-238、-857、-863位点，IL-10启动子-1082、-592位点及黏病毒抵抗蛋白A（MxA）启动子-88位点单核苷酸多态性（SNP）的关系。方法：305例CHB患者IFNα-1b治疗12个月，随访至停药后6个月判断疗效，分为持续应答（SR）和非持续应答（NSR）组。应用PCR及限制性片段长度多态性方法，检测TNF-α-238、-857、-863和IL-10—592、-1082及MxA-88位点的SNP。判断SNP与IFN疗效的关系。结果：本研究失访43例。262例CHB患者IFNα-1b疗效SR为50例，占19．1％；NSR212例，占80．9％。在MxA-88位点，GT型分别与GG型、TT型患者IFN疗效比较，差异均有统计学意义（x^2=20．119，OR：5．302，95％CI：2．458～11．433，P〈0．01；x^2=13．071，OR：4．110，95％CI：1．858～9．092，P〈0．01）。在TNF—α-863位点，CC型分别与CA型、AA型患者疗效比较，差异均有统计学意义（x^2=29．628，0R：7．578，95％CI：3．444～16．672，P〈0．01；x^2=13．543，0R：4．513，95％CI：1．966～10．357，P〈0．01）。在TNF-α-857位点，CC与CT型患者疗效比较，差异有统计学意义（x^2=12．927，OR：0．293，95％CI：0．146～0．586，P〈0．01）。在IL广10-592位点，AA与CC型患者疗效比较，差异有统计学意义（x^2=8．984，OR：3．380，95％CI：1．484～7．697，P〈0．01）。结论：MxA-88位点为GT杂合型，TNF—α-863 CC纯合型，IL-10—592AA纯合型的CHB患者对IFNα-1b治疗反应好，可作为预测IFN疗效的参考指标之一。     

The immunoneuroendocrine role of melatonin

Abstract: A tight, physiological link between the pineal gland and the immune system is emerging from a series of experimental studies. This link might reflect the evolutionary connection between self-recognition and reproduction. Pinealectomy or other experimental methods which inhibit melatonin synthesis and secretion induce a state of immunodepression which is counteracted by melatonin. In general, melatonin seems to have an immunoenhancing effect that is particularly apparent in immunodepressive states. The negative effect of acute stress or immunosuppressive pharmacological treatments on various immune parameters are counteracted by melatonin. It seems important to note that one of the main targets of melatonin is the thymus, i.e., the central organ of the immune system. The clinical use of melatonin as an immunotherapeutic agent seems promising in primary and secondary immunodeficiencies as well as in cancer immunotherapy. The immunoenhancing action of melatonin seems to be mediated by T-helper cell-derived opioid peptides as well as by lymphokines and, perhaps, by pituitary hormones. Melatonin-induced-immuno-opioids (MHO) and lymphokines imply the presence of specific binding sites or melatonin receptors on cells of the immune system. On the other hand, lymphokines such as -γ-interferon and interleukin-2 as well as thymic hormones can modulate the synthesis of melatonin in the pineal gland. The pineal gland might thus be viewed as the crux of a sophisticated immunoneuroendocrine network which functions as an unconscious, diffuse sensory organ.     

Response of rat pineal melatonin to calcium, magnesium, and lithium is circadian stage dependent

Abstract: Herein we documented the response of pineal melatonin production to electrolytes known to be effective on pineal function in view of a possible circadian stage dependence. We studied the release of melatonin by perifused rat pineal glands at 2 different circadian stages corresponding to the middle of the light and dark periods, i.e., respectively, 7 and 19 HALO (Hours After Light Onset, L:D = 12:12). The initial efflux rates were, as expected, much higher in the perifusates of glands removed from rats sacrificed during the dark phase than of those removed during the light phase. After 3 hr of perifusion, melatonin release reached similar levels which were found constant up to the 8th hr of perifusion, whatever the circadian stage. Perifusion of the glands with physiological concentrations for the rat of calcium (5.2 mmol/1) and magnesium (1.34 mmol/1) resulted in a stimulatory effect on the pineal glands removed from rats sacrificed in the middle of the dark period (19 HALO), whereas no effects were observed on the pineal glands removed from rats sacrificed during the light (7 HALO). Lithium (0.28 and 0.55 mmol/1) was ineffective on melatonin release in pineal glands removed 7 and 19 HALO. Our results show differences in the initial efflux rates of melatonin and in the response of perifused pineal glands to calcium and magnesium according to the circadian stage.     

Changes in connexin43, the gap junction protein of astrocytes, during development of the rat pineal gland

Abstract: The abundance of gap junctions between rat pineal astrocytes formed by connexin43 (Cx43) was studied during development. Levels and distribution of Cx43 were measured by immunoblotting and indirect immunofluorescence, respectively. The amount of Cx43 in cells located within the gland was low until about the 7th postnatal day and increased to adult values between the 14th and 21st days postpartum. Although astrocytes, recognized by their vimentin immunoreactivity, were scarce before birth, they were abundant by the 7th postnatal day suggesting that the low levels of Cx43 found at this age corresponded to a low expression of this protein. Localization of the immunoreactivity to Cx43 and vimentin showed a close correlation, indicating that mature or immature pineal astrocytes form gap junctions made of Cx43. Since Cx43 levels attained their adult values at about the time the innervation and the functional state of the gland reached maturity (2–3 weeks after birth), it is proposed that astrocyte gap junctions are involved in the function of the adult rat pineal gland.     

Conservative management of perforated duodenal diverticulum： A case report and review of the literature

Duodenal diverticula are a relatively common condition. They are asymptomatic, unless they become complicated, with perforation being the rarest but most severe complication. Surgical treatment is the most frequently performed approach. We report the case of a patient with a perforated duodenal diverticulum, which was diagnosed early and treated conservatively with antibiotics and percutaneous drainage of secondary retroperitoneal abscesses. We suggest this method could be an acceptable option for the management of similar cases, provided that the patient is in good general condition and without septic signs.     

Presence of immunoreactive growth hormone and prolactin in the ovine pineal gland

Abstract: The use of antisera raised against bovine growth hormone (GH) and ovine prolactin (PRL) enabled the detection of related immunoreactive (ir) sequences of proteins in ovine pineal tissue. The isolation of PRL-like ir-material was accomplished using a 0.25 M ammonium sulphate (pH 5.5) extraction followed by ethanol precipitation, whereas the resulting 2.0 M ammonium sulphate (pH 7.0) precipitate contained a GH-like immunoreactivity. Gel chromatography of the GH-like immunoreactivity (Sephadex G-100) indicated the presence of several GH-like fragments ranging in the M_r range of 7,000 to 55,000. Analyses of the PRL-like ir-material found in pineal tissue on HPLC using a TSK 545-DEAE column led to the resolution into a single peak of immunoreactivity. A single peak of activity was also observed following chromatofocusing and hydrophobic interaction chromatography of the ir-peak from the TSK 545-DEAE column. The PRL-like ir-material inhibited the binding of [¹²⁵I]ovine PRL-S14 to anti-ovine PRL antibodies without showing an affinity for binding to anti-rat PRL or anti-bovine GH antibodies. Scatchard analysis of the binding of pineal PRL-like ir-material and pituitary ovine PRL-S14 to liver membranes from day-20 pregnant rats revealed similar affinity constants (K_a of 4.7 ± 0.2 × 10⁹ M^-1). In addition, the replication of Nb 2 Node rat lymphoma cells was stimulated by pineal PRL-like ir-material, an effect known to be specific for lactogenic hormones. The pineal PRL-like immunoreactivity appeared on sodium dodecyl sulfate polyacrylamide gels as a single major band of M_r 24,000. The functional status of PRL-and GH-like ir-material in the ovine pineal remains to be determined, but evidence is presented that the overall protein synthesis rate of the rat pineal responded to circulating concentrations of PRL.     

Microbiology of human immunodeficiency virus anorectal disease

PURPOSE: Individuals who are seropositive for the human immunodeficiency virus are at high risk for opportunistic infection and anorectal disorders. Little prospective information is available regarding anorectal pathogens in these patients. METHODS: One hundred sixty-three HIV-seropositive patients presented to the colorectal clinic between 1989 and 1992. Forty-seven (29 percent) patients were thought to have an infectious process and were prospectively studied using a standardized multiculture protocol. RESULTS: Mean age was 33 (range, 19–59) years. All were male; high-risk behavior accounted for 87 percent of HIV transmissions. Presenting complaints included anorectal pain (79 percent), pus per anum (28 percent), and blood per anum (26 percent). Examination revealed perianal tenderness (60 percent), condyloma (38 percent), perianal ulcers (38 percent), and anal fissures (34 percent). Sixty-six sets of cultures were performed; 28 patients had one set, 15 had two sets, and 4 had three sets. Thirty-two of these 47 patients (68 percent) had positive cultures including herpes (50 percent), cytomegalovirus (25 percent),Neisseria gonorrhoeae (16 percent), chlamydia (16 percent), acidfast bacilli (2 percent), and others (9 percent). Six of 32 patients with positive cultures had more than one organism cultured. Sixteen (50 percent) patients with positive cultures were treated medically, 8 (25 percent) were treated surgically and 8 (25 percent) were treated with both modalities. Sixty-one procedures were performed on 17 patients for condylomata. Eighteen patients had 20 procedures for abscesses, 50 percent of whom had positive cultures for other than common bowel flora; all improved. Fourteen patients underwent 33 procedures for perianal fistulas.Mycobacterium fortuitum was cultured from one patient who required 13 procedures for abscesses and fistulas. Forty-five (96 percent) patients were followed for an average of 12.5 months ±2.9 SEM (range, 1–94 months). Symptoms were improved or resolved in 22 of 32 (69 percent) patients with positive cultures and in 11 of 13 (84 percent) with negative cultures. CONCLUSIONS: Specific pathogens may often be identified in human immunodeficiency virus-seropositive patients with anorectal disorders if aggressively sought. Although patients without specific pathogens identified may be expected to improve with planned empiric treatment, positive identification allows more directed therapy.