1.采用分期辨证,取代分型辨证 自从"中医辨证与西医辨病相结合"的诊疗模式确定以来,现代肝脏病学的临床分类体系已为中西医结合肝病从业者所遵从.但是,由姜春华教授和沈自尹院士所创立的"分型辨证"模式,却受到了极大的挑战.笔者在多篇文章和著作中专门论述过这一模式的内在缺陷,并大力推荐"分期辨证"取代"分型辨证".这是"证"的定义所决定的,即机体在疾病发展过程中某一阶段病理反映的高度概括.

按:王伯祥老师辞世后,大家提出办一期纪念专刊.组稿过程中,《中西医结合肝病杂志》编辑部希望王老的学生们完成一篇"师说"的纪念文章.想到2018 年12月,《中西医结合肝病杂志》编委会召开之际,我到王老师府上看望他,其中与老师谈到中医肝病学术的未来发展,今就记忆所及,加工整理如下.

本文回顾了王伯祥教授治疗肝病尤重瘀血的学术思想和善用活血化瘀药物的临床经验,总结了该团队在"肝病血瘀证"临床和实验研究的成果,阐述了该团队在此基础上构建"毒损肝络"假说的具体内容和临床应用,提出以分期辨证取代分型辨证诊疗体系的构想.

目的 探索维生素D水平对替诺福韦治疗慢性乙型肝炎24周病毒学应答的影响.方法 纳入2017年11月至2019年11月期间深圳市第三人民医院肝病医学中心收治的患者共115例,收集基线临床资料,包括人口学信息、病毒学指标、血常规、肝功能、凝血功能以及维生素D水平,根据治疗24周乙肝病毒定量结果分为阳性组和阴性组,通过单因素及多因素Logistic分析得出维生素D与病毒学应答的关系,将性别、HBeAg状态、HBV DNA水平作为分层因素进行亚组分析.结果 HBV DNA阳性组基线25(OH)D水平明显低于HBV DNA阴性组(P=0.002),多因素分析显示,24周HBV DNA转阴率OR值随着25(OH)D水平逐渐增加,亚组分析中,24周HBV DNA转阴率OR值均随着25(OH)D水平的升高而增加.结论 25(OH)D水平是影响替诺福韦治疗慢性乙型肝炎24周病毒学应答的独立影响因素.

目的:分析非结核分枝杆菌肺病(NTM-PD)治疗结局并探索其影响因素.方法:收集深圳市第三人民医院2012年11月至2017年7月确诊为NTM-PD的242例患者的年龄、性别、基础疾病、症状、影像学改变、药敏试验、治疗随访情况资料,按治疗结局分为治愈组和非治愈组,采用SPSS 22.0统计软件分析影响治疗结局的因素.结果:非结核分枝杆菌耐药率高,总体治愈率为58.7％,Logisitc回归分析显示发热[OR=3.067,95％CI(1.223,7.688)]、乙胺丁醇(EMB)耐药[OR=2.741,95％CI(1.448,5.191)]、支气管扩张[OR=2.781,95％CI(1.469,5.263)]及未规律治疗[OR=3.700,95％CI(1.859,7.364)]是影响治疗结局的危险因素.结论:发热、EMB耐药可能增加治疗难度;NTM-PD治愈率低,与支气管扩张、未规律治疗相关.

目的 中西医结合治疗乙型肝炎病毒(HBV)感染的临床疗效评价.方法 196例HBV感染患者作为研究对象,采用数字随机法分为联合组和对照组,各88例.对照组采用西医临床治疗,联合组在对照组基础上联合中医治疗,对比两组的乙型肝炎e抗原(HBeAg)转阴情况、乙肝病毒的脱氧核糖核酸(HBV-DNA)转阴情况、满意度以及治疗前后白介素-2(IL-2)和γ-干扰素(IFN-γ).结果 联合组HBeAg转阴23例,转阴率为26.14％;对照组转阴8例,转阴率为9.09％;两组HBeAg转阴率对比差异具有统计学意义(χ2=8.810,P=0.003<0.05).联合组HBV-DNA转阴33例,转阴率为37.50％;对照组HBV-DNA转阴14例,转阴率为15.91％;两组HBV-DNA转阴率对比差异具有统计学意义(χ2=10.479,P=0.001<0.05).治疗前,两组IL-2、IFN-γ对比差异无统计学意义(P>0.05);治疗后,两组IL-2、IFN-γ水平均高于治疗前,且联合组高于对照组,差异具有统计学意义(P<0.05).两组满意度对比差异具有统计学意义(χ2=5.274,P=0.022<0.05).结论 中西医结合治疗HBV感染具有良好的临床疗效,能有效缓解患者临床症状,改善患者预后,值得推广应用.

Objective:To study the correlation between the IL-28 B gene polymorphism and Traditional Chinese Medicine syudrome,liver biochemical characteristics and APRI in patients with chronic hepatitis C.Methods:Pyrosequencing method was used to test genotype IL28B gene rs12979860 SNP and completed the liver biochemistry test,blood routine test,ultrasound examinatio in 105 cases of naive chronic HCVpatients visited our hospital.Then the TCM syndrome types were classified and the distribution of TCM Syndromes and their correlation with biochemical characteristics and APRI score were compared.Results:There were 90 cases CC type of IL28B gene in 105 chronic HCV patients visited our hospital.,the liver and kidney yin deficiency was the most common TCM syndrome in these patients(37 cases,41.1％),AST and APRI score in this TCM syndrome were significantly higher than other syndromes,and the percentage of liver cirrhosis was the highest in this syndrome.(P ＜ 0.05).Conclusion:The liver biochemical abnormalities and APRI score are most obvious in the liver and kidney Yin deficiency syndrome of IL28B allele for CC in patients with chroinc HCV,so the patients in liver and kidney Yin deficiency syndrome has the higher potential risk to cirrhosis.

Objective To explore the feasibility of compound tonic Chinese herbal formula as the adjuvant on strengthening the immunogenicity of influenza virus-like particles( VLPs)vaccine. Methods Forty-two BALB/C mice were randomized into 7 groups,6 mice in each one,i. e. VLPs group,VLPs+qi tonic formula group,VLPs+blood nourishing formula group,VLPs+yin nourishing formula group,VLPs+yang tonic formula group,VLPs+cholera toxin group and PBS negative control group. Separately,on 0 and 14th days,VLPs were used via nasal feeding. From the 1st to the 13th day,Chinese herbal medicine was used in lavage in the compound tonic Chinese herbal formula groups. VLPs were vaccinated at 10μg per time,Chi-nese herbal adjuvant agents were at 200 mg/kg and cholera toxin was at 1 μg vas nasal vaccination. The pe-ripheral blood was collected before vaccination,on the 13th and 28th days after vaccination. BALF was collect-ed on the 28th day. The levels of IgG in peripheral serum and IgA in BALF were determined and further,the neutralizing antibody titer in peripheral serum was determined. Results 1. Chinese tonic herbal medicine en-hanced IgG fluid immune response in peripheral blood,in which,the qi tonic formula and yin nourishing for-mula rapidly increased IgG level at early stage(P﹤0. 01)and the qi tonic formula further persistently in-creased IgG level(P﹤0. 01). 2. The qi tonic formula,blood nourishing formula and yin nourishing formula significantly increased IgA level induced by VLPs(P﹤0. 01),of them,the increase was the most apparent&nbsp;with the qi tonic formula. 3. No matter at the early or later stage of immune response,the qi tonic formula and yin nourishing formal all significantly improved the neutralizing antibody titer in peripheral blood in the mice induced by VLPs(P ﹤0. 01). Conclusion Qi tonic formula and yin nourishing formula significantly in-crease peripheral fluid immune response and lung local mucous immune response induced by VLPs. The com-bination of the formula for qi tonic and yin nourishing is anticipant to be a new Chinese herbal adjuvant and enhance VLPs immunogenicity.

OBJECTIVE:To investigate whether Chinese medicine (CM) recipes could ameliorate H5N1 influenza virus infection in BALB/c mice model.METHODS:BALB/c mice were orally administrated with 5 CM recipes (removing toxin, tonifying qi, cooling blood, laxation, and compound recipes), oseltamivir, or saline solution respectively for 5 consecutive days after the infection of H5N1 influenza virus. Series of indices were employed to evaluate the amelioration of the 5 CM recipes on infection, including clinical assessment, gross observation, histopathologic findings, cytokine levels and viral burden in the lungs.RESULTS:Two CM recipes (cooling blood and compound recipes) could postpone the death period of the mice infected with high-dose H5N1 influenza virus (P< 0.05). And for the mice infected with low-dose H5N1 influenza virus, CM recipes could significantly reduce the mortality and inhibit viral proliferation in the lungs as compared with the control group (P<0.05). There was no significant difference in lung coefficients between the treatment and the control groups, but histopathological findings in the lungs were improved in CM recipes groups compared to control group findings. A transient increase was observed in pro-inflammatory and anti-inflammatory cytokines during the first 6 days of infection. The levels of interleukin (IL)-12p40 and interferon-gamma of the treatment groups were significantly lower than that of the control group at day 3 post-infection (P<0.05), while only compound recipe were significantly lower in level of tumor necrosis factor alpha than the control (P<0.05). The level of IL-10 of the control was higher than others, and the differences between the control and cooling blood, removing toxin recipes were significant (P<0.05).CONCLUSIONS:Results of this study suggested the potentials of the CM recipes in ameliorating influenza virus infection by suppressing viral proliferations, improving histopathological lesions, and inhibition of over expression of inflammatory cytokines.

Objective To explore the discrepancy between IL-1β and IL-1Ra expression in CD14+ monocytes of different TCM syndrome types of pulmonary tuberculosis patients. Methods Totally 104 pulmonary tuberculosis patients were recruited,the patients were divided into Lung-Yin deficiency( n=33),Yin-Deficiency and Fire-Hyperactivity(n=36),and Qi and Yin deficiency(n=35). The density gradient centrifugation was used to separate PBMCs from peripheral blood, then CD14+ monocytes were purified after incubation with magnetic beads. Monocytes were stimulated with or without Mtb lysate for 12&nbsp;hours,the supernatant was collected to detect IL-1β and IL-1Ra levels using ELISA,and total RNA was purified from cells and gene expression of IL1B and IL1RN was determined with real-time qPCR assay. Finally,the discrepancy was compared with ANOVA analysis. Results (1) The expression of IL1B and IL1RN could be detected by real-time quantitative PCR assay with gene specific primers. (2) After Mtb stimulation,IL-1β level was the highest in Lung-Yin deficiency group compared to Yin-Deficiency and Fire-Hyperactivity group and Qi and Yin deficiency group(P<0. 05、P<0. 01), and it was higher in Yin-Deficiency and Fire-Hyperactivity group than that in Qi and Yin deficiency group(P<0. 05). There was similar pattern observed in IL1B mRNA level, and IL1B level decreased in the order of Lung-Yin deficiency, Yin-Deficiency and Fire-Hyperactivity, and Qi and Yin deficiency(P<0. 05、0. 01). (3)IL-1 Ra expression increased significantly after Mtb stimulation, it was the highest in Qi and Yin deficiency group compared to other two groups(P<0. 05), and there was no difference between Yin-Deficiency and Fire-Hyperactivity group and Qi and Yin deficiency group(P>0. 05). Similar pattern was also observed in IL1RN level,which Qi and Yin deficiency group had the highest level(P<0. 05, P<0. 01). (4)After Mtb stimulation, IL-1β/IL-1Ra ratio increased in Lung-Yin deficiency and Yin-Deficiency and Fire-Hyperactivity groups,which was higher than that in Qi and Yin deficiency(P<0. 01, P<0. 05). Conclusion Expression of IL-1β and IL-1Ra was associated with TCM pathogenesis evolution of pulmonary TB patients, which could be considered as biomarkers of microcosmic syndrome differentiation and effectively therapeutic targets.

目的:探讨肝硬化难治性腹水患者的中医证候特征,为肝硬化难治性腹水的中医治疗提供辨证施治依据.方法:采用前瞻性流行病学调查方法,提取94例肝硬化难治性腹水患者的中医症状、体征、舌、脉等四诊信息,对患者进行中医辨证,进行统计分析这类患者的证候学特点.结果:肝硬化难治性腹水患者常见的中医证候依次为:脾虚湿阻证(80.8％),血瘀证(71.3％),湿热蕴结证(55.3％),肝气郁结证(44.7％),肝肾阴虚证(9.6％),脾肾阳虚证(4.3％).其中同一患者常有几个证候重叠.结论:肝硬化难治性腹水患者的中医证候较少见单一证候,兼夹证多见.脾虚湿阻兼夹瘀热的复合证型,是肝硬化难治性腹水的最特征性证候.

Objective To explore the regulating effects of tonic herbal compound on the IL-17A expression induced by influenza virus-like particles in peripheral blood. Methods 42 BALB/C mice were divided into 7 groups(n= 6 for each group),involving single VLPs group,VLPs combined Yiqi recipe, VLPs combined Buxue recipe,VLPs combined Ziyin recipe,and VLPs combined Zhuangyang recipe group, VLPs combined cholera toxin group was considered as positive control,and PBS only group was considered as blank control. The mice were immunized with 10μg VLPs each time through intranasal administration in day 0 and 14,and herbs were intragastric administration with 200 mg/Kg from first day to day 13. The blood was collected to make serum and PBMCs at pre-immunization and day 13 and day 28 after immunization. The IL-17A level in serum was determined by ELISA,and spot forming cells( SFCs) of IL-17A were determined using Elispot assay. Results (1)No matter in the early stage or late stage,influenza VLPs could induce IL-17A expression in serum, especially, Yiqi recipe could increase IL-17A level compared to single VLPs group(P<0. 01). (2)Elispot technology can effectively detect the secretion of IL-17A positive cells in PBMCs. The SFCs of IL-17A was much higher in the Yiqi recipe and Ziyin recipe (P<0. 01),especially at day 28 after immunization. Conclusion Yiqi recipe and Ziyin recipe can effectively increase IL-17A expression induced by influenza VLPs,which could be considered as new type adjuvants for influenza vaccine.

目的:研究赤芍承气汤对内毒素二次打击后急性肝损伤大鼠细胞因子的影响,阐述其对肝衰竭的防治机制.方法:将75只SD大鼠随机分为正常组(A),模型组(B),赤芍承气汤低(C)、中(D)、高(E)剂量组,培菲康组(F),造模前3天开始灌胃,末次灌胃1h后,除正常组外其余组大鼠通过D-氨基半乳糖腹腔注射24h后成肝衰竭模型对SD大鼠制造第一次打击,之后给予赤芍承气汤或培菲康治疗,再腹腔注入脂多糖(LPS,内毒素的主要成分)作为第二次打击.观察大鼠行为学变化,二次打击后2h、8h处死大鼠,测定各组大鼠血清天门冬氨酸转氨酶(AST)、丙氨酸转氨酶(ALT)、总胆红素(TBil)、内毒素(ET)水平、血浆肿瘤坏死因子-α(TNF-α)、白介素(IL)-6、IL-1β,观察肝组织病理学.结果:二次打击致各组大鼠ALT、AST、TBil、ET、TNF-α、IL-6、IL-1β水平明显高于正常组,肝脏内炎细胞浸润、坏死明显,药物干预组大鼠相关指标较模型组明显降低(P＜0.05),肝组织病变减轻.结论:赤芍承气汤对内毒素二次打击急性肝损伤大鼠具有防治作用,该方能减轻其肝脏的损伤,改善肝脏功能,提高其大鼠生存率,其疗效机制可能与减轻内毒素血症从而切断了部分内毒素的信号传导通路,有效抑制炎症信号通路的激活,减轻TNF-α等炎症介质的释放进而减轻肝损伤有关.

Objective To produce influenza virus-like particles(VLPs) with drosophila S2 cell line and identify its bioactivity.Methods The persistent expression vector sof pAc-V5-HA,NA and M1 suitable for drosophila S2 cell line were constructed at the earlier stage,which were co-transfected into S2 cell with pCoblast vector.Then we screened monoclonal S2 cell lines which integrated HA,NA and M1 genes through blasticindin combined with limited dilution methods.We used PCR assay to identify whether HA,NA and M 1 genes were successfully integrated into genome,and used Western-Blot assay to identify whether there was VLPs in the supernatant.The HA activity was evaluated with hemagglutination test,and NA activity was reflected with the ability to hydrolyze sialic acid.Results The HA,NA and M1 genes were successfully amplified from the monoclonal S2 cell using PCR assay.The VLPs integrated HA,NA and M1 protein was identified with Western-Blot assay.The hemagglutination test confirmed influenza VLPs could agglutinate chicken red blood cells with titer of 1 ∶ 160.The NA could dose-dependent hydrolyze sialic acid,and no significance was observed at the high dose(250 ng,500 ng)between wild type virus and VLPs.Conclusion We successfully established the monoclonal S2 cell line which integrated influenza HA,NA and M 1 genes,and it could persistently produce influenza VLPs with perfect HA and NA activity.VLPs could be considered as effective influenza vaccine candidate in the future.

患者的一封来信 尊敬的聂广主任: 我想给您写这封信已经很久了,提笔祝您工作顺利,心想事成. 两个月来,我一直在考虑抗病毒治疗一事.但决心一下,眼泪就止不住地流出来了.迟迟不敢踏上抗病毒治疗这条前途未卜又没有归期的“贼船”,主要出于三方面原因:

Abstract Objective To realize the characteristics of “zheng” differentiation-treatment for hand, foot and mouth disease (HFMD), a new methodology of syndrome differentiation for different stages of HFMD has been explored. Methods Total of 2 325 cases with HFMD were recorded by distributing them into exterior syndrome stage, interior syndrome stage, severe syndrome stage and recovered syndrome stage, respectively, and the main symptoms and subsidiary symptoms of different stages of HFMD have been observed. The major and minor pathogenesis of HFMD in different stages were obtained, and compared with the “2010 Guideline for the Diagnosis and Treatment of HFMD”. Results It was found that the major pathogenesis of exterior stage was defined as “the invation of the wenevil to the defender of the body with the collaterals got involved”, and the minor as “qi deficiency”; in interior stage, “the fury of Gan-Yang” was the main pathogenesis, and “qi in chaos and qi deficiency” was the minor; in severe syndrome stage, “the damage of heart, liver and lung” was the main pathogenesis, and “qi in chaos” was the minor; and the pathogenesis of recovered stage was “qi-yin deficiency”. Compared with the “2010 Guideline for the Diagnosis and Treatment of HFMD”, it showed that “the obstruction of the fei-pi qi by the mixture of shi-re evil” and “the mixture of shi-re” in vivo was quite difficult to be explained in completely different context in the general situation; in the severe stage, the TCM clinical characteristics of syndrome differentiation might lose; in the early acute severe cases, the phenomenon that xin-yang and fei-qi almost ran out was difficult to be observed, then, the line between the severe and the acute severe became vague. Conclusions The theory of syndrome differentiation by stages of HFMD was reasonable in the actual situation of clinical description on HFMD which was expected to be further tested and widely applied in the “zheng” differentiation-treatment of HFMD in the future.

目的 通过对狂犬病临床表现的回顾性研究,初步探讨狂犬病不同分期中医证候要素的分布情况并总结病因病机,为中医临床辨证提供可能的参考.方法 收集符合狂犬病诊断的病例36例,按照超早期、早期、中期、晚期整理分析其症状、体征.采用描述性统计,初步探讨不同分期中医症状及证候要素的分布情况.结果 狂犬病超早期以毒伏脉络为主;早期基本病机为风毒内动出表,次要病机为正气亏虚、瘀阻脉络;中期以瘀热入络、肝风内动、痰蒙心窍为主,次要病机为气阴两虚;晚期病机为热毒互结,痰瘀阻闭,阳脱阴竭.结论 根据狂犬病临床表现确定中医"证素",结合病因病机及伏气温病理论分期辨证更能体现狂犬病的发生、发展过程、临床表现的时空特点,为中医药防治狂犬病提供可能的依据.

The quality of turmeric products is directly based on the amount of curcumin. In this study, the Raman spectrum of curcumin was obtained to analyze curcuminoids in turmeric products quantitatively, particularly turmeric roots in powder form. The Raman band of sodium sulfite was used as an internal standard to remove the effect of several factors such as laser power and other instrumental parameters. Band intensity ratios ( I 1,601 / I 989 and I 1,630 / I 989 ) were calculated for linear fitting. A calibration curve was generated by plotting the band ratio against sample concentration. Results showed that the band ratio between the Raman intensity of the samples and the internal standard exhibited good linearity with analyte concentration. This model was subsequently used to quantitatively analyze curcumin in different layers of turmeric roots, successfully confirming the ranges of curcumin weight ratio. Thus, the proposed method could be a convenient alternative to standard procedures used to determine the quality of turmeric roots.

In the present study, Raman spectral characteristics of methemoglobin (MetHb) induced by sodium nitrite (NaNO2) were investigated. Hemoglobin (Hb) was oxidated to MetHb with NaNO2, the Raman spectral specific changes of MetHb was studied by determining the Raman spectral changes of methemoglobin/total hemoglobin of different ratios, and the Raman intensities of methemoglobin/total hemoglobin of different ratios at 1 586, 1 605 and 1 637 cm(-1) were linearly fitted to realize its quantitative detection. The results show that the completely oxidized MetHb can be obtained when the molar ratio of NaNO2 to Hb is 3.5 : 1 whose Raman characteristic peaks are at around 499, 1 340, 1 562 and 1 622 cm(-1), and that the linear fitting correlation coefficients R2 of the Raman intensities of methemoglobin/total hemoglobin of different ratios at 1 586, 1 605 and 1 637 cm(-1) are 0.972 84, 0.997 97 and 0.991 26 respectively, which shows a good linear relationship. This study indicates that the Raman spectrums of MetHb induced by NaNO2 have characteristic differences when compared with normal Hb, that the locations and intensities of Raman characteristic peaks change correspondingly with the alterations of the ratios of methemoglobin/total hemoglobin, and that there are linear correlations between the ratios and their corresponding Raman intensities, which would provide theoretical bases for the clinical Raman spectral detection and quantitative study of methemoglobinemia.

BACKGROUND:Host genetic variations may contribute to disease susceptibility of influenza. IL-1A and IL-1B are important inflammatory cytokines that mediate the inflammation and initiate the immune response against virus infection. In this study, we investigated the relationship between single-nucleotide polymorphisms (SNPs) of Interleukin-1A (IL-1A) and Interleukin-1B (IL-1B) and the susceptibility to 2009 pandemic A/H1N1 influenza (A(H1N1)pdm09). 167 patients whom were confirmed with A(H1N1)pdm09 and 192 healthy controls were included in this study. Four SNPs (rs1304037, rs16347, rs17561, rs2071373) in IL1A gene and three SNPs (rs1143623, rs3917345, rs1143627) in IL1B gene were genotyped by using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry platform, and the associations of the genetic variants of IL-1 with susceptibility to A(H1N1)pdm09 were then assessed.RESULTS:The polymorphisms of rs17561 in IL1A gene and rs1143627 in IL1B gene were found to be associated with susceptibility to A(H1N1)pdm09 with P values of 0.003 (OR 2.08, 95% CI 1.27-3.41) and 0.002 (OR 1.62 , 95% CI 1.20-2.18), respectively. However, no significant difference in allelic frequency was observed for other SNPs between cases and controls.CONCLUSIONS:This study provides a new insight into pathogenesis of A(H1N1)pdm09, suggesting that genetic variants of IL-1A and IL-1B may exert a substantial impact on the susceptibility of A(H1N1)pdm09 virus infection.