Discrimination of Two Cultivars of Alpinia Officinarum Hance Using an Electronic Nose and Gas Chromatography-Mass Spectrometry Coupled with Chemometrics.

Background: Alpinia officinarum Hance is both an herbal medicine and a condiment, and generally has different cultivars such as Zhutou galangal and Fengwo galangal. The appearance of these A. officinarum cultivars is similar, but their chemical composition and quality are different. It is therefore important to discriminate between different A. officinarum plants to ensure the consistency of the efficacy of the medicine. Therefore, we used an electronic nose (E-nose) to explore the differences in odor information between the two cultivars for fast and robust discrimination. Methods: Odor and volatile components of all A. officinarum samples were detected by the E-nose and gas chromatography-mass spectrometry (GC-MS), respectively. The E-nose sensors and GC-MS data were analyzed respectively by principal component analysis (PCA), the correlation between E-nose sensors and GC-MS data were analyzed by partial least squares (PLS). Results: It was found that Zhutou galangal and Fengwo galangal can be discriminated by combining the E-nose with PCA, and the E-nose sensors S2, S6, S7, S9 were important sensors for distinguishing different cultivars of A. officinarum. A total of 56 volatile components of A. officinarum were identified by the GC-MS analysis, and the composition and content of the volatile components from the two different A. officinarum cultivars were different, in particular the relative contents of 1,8-cineole and α-farnesene. The classification result by PCA analysis based on GC-MS data was consistent with the E-nose results. The PLS analysis demonstrated that the volatile terpene, alcohol and ester components primarily interacted with the sensors S2 and S7, indicating that particular E-nose sensors were highly correlated with some aroma constituents. Conclusions: Combined with advanced chemometrics, the E-nose detection technology can discriminate two cultivars of A. officinarum, with GC-MS providing support to determine the material basis of the E-nose sensors' response.

The role of autophagy in THP-1 macrophages resistance to HIV- vpr-induced apoptosis.

Macrophages are resistant to cell death and are one of HIV reservoirs. HIV viral protein Vpr has the potential to promote infection of and survival of macrophages, which could be a highly significant factor in the development and/or maintenance of macrophage viral reservoirs. However, the impact of vpr on macrophages resistance to apoptosis is yet to be comprehended. Autophagy is a cell survival mechanism under stress state. In this study, we investigated whether autophagy is involved in macrophages resistant to vpr-induced apoptosis. Using the THP1 macrophages, we studied the interconnection between macrophages resistance to apoptosis and autophagy. We found that vpr is able to trigger autophagy in transfected THP-1 macrophages confirmed by electron microscopy (EM) and western blot analysis, and inhibition of autophagy with 3MA increased vpr-induced apoptosis. The results indicate that autophagy may be responsible for maintenance of macrophage HIV reservoirs.

Identification of Chinese Herbal Medicines with Electronic Nose Technology: Applications and Challenges.

This paper provides a review of the most recent works in machine olfaction as applied to the identification of Chinese Herbal Medicines (CHMs). Due to the wide variety of CHMs, the complexity of growing sources and the diverse specifications of herb components, the quality control of CHMs is a challenging issue. Much research has demonstrated that an electronic nose (E-nose) as an advanced machine olfaction system, can overcome this challenge through identification of the complex odors of CHMs. E-nose technology, with better usability, high sensitivity, real-time detection and non-destructive features has shown better performance in comparison with other analytical techniques such as gas chromatography-mass spectrometry (GC-MS). Although there has been immense development of E-nose techniques in other applications, there are limited reports on the application of E-noses for the quality control of CHMs. The aim of current study is to review practical implementation and advantages of E-noses for robust and effective odor identification of CHMs. It covers the use of E-nose technology to study the effects of growing regions, identification methods, production procedures and storage time on CHMs. Moreover, the challenges and applications of E-nose for CHM identification are investigated. Based on the advancement in E-nose technology, odor may become a new quantitative index for quality control of CHMs and drug discovery. It was also found that more research could be done in the area of odor standardization and odor reproduction for remote sensing.

Hepatitis B virus mutation may play a role in hepatocellular carcinoma recurrence: A systematic review and meta-regression analysis.

BACKGROUND AND AIMS: A number of studies have confirmed that antiviral therapy with nucleotide analogs (NAs) can improve the prognosis of hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) after curative therapy. However, what factors affected the prognosis of HBV-HCC after removal of the primary tumor and inhibition of HBV replication? A meta-regression analysis was conducted to explore the prognostic factor for this subgroup of patients. METHODS: MEDLINE, EMBASE, Web of Science, and Cochrane library were searched from January 1995 to February 2014 for clinical trials evaluating the effect of NAs on the prognosis of HBV-HCC after curative therapy. Data were extracted for host, viral, and intervention information. Single-arm meta-analysis was performed to assess overall survival (OS) rates and HCC recurrence. Meta-regression analysis was carried out to explore risk factors for 1-year OS rate and HCC recurrence for HBV-HCC patients after curative therapy and antiviral therapy. RESULTS: Fourteen observational studies with 1284 patients met the inclusion criteria. Influential factors for prognosis of HCC were mainly baseline HBeAg positivity, cirrhotic stage, advanced Tumor-Node-Metastasis (TNM) stage, macrovascular invasion, and antiviral agent type. The 1-year OS rate decreased by more than four times (coefficient -4.45, P<0.001) and the 1-year HCC recurrence increased by more than one time (coefficient 1.20, P=0.003) when lamivudine was chosen for HCC after curative therapy, relative to entecavir for HCC. CONCLUSIONS: HBV mutation may play a role in HCC recurrence. Entecavir or tenofovir, a high genetic barrier to resistance, should be recommended for HBV-HCC patients.

Clinical Characteristics of Non-B and Non-C Biopsy-Proven Primary Liver Cancers in an HBV- Endemic Area: A Retrospective Study.

Objective: To explore the distribution of probable causes and clinical characteristics of non-B and non-C (NBNC) primary liver cancer (PLC) patients in the HBV-endemic region. Methods: A total of 86 individuals with biopsy-proven NBNC-PLC were enrolled. NBNC-PLC patients were defined as negative for both anti-HCV antibodies and five serum hepatitis B markers. Patients' characteristics were collected from medical records. Results: Among them, most of the NBNC-PLC patients had intrahepatic cholangiocarcinoma (ICC) (81.4%), and 12.8% had hepatocellular carcinoma (HCC). The NBNC ICC group had more platelet count, GGT, and CA199 levels; approximately two-thirds were female, and it was more often present in patients with biliary inflammatory diseases, especially intrahepatic biliary lithiasis. The NBNC HCC group was older and had a higher proportion of dyslipidemia, obesity, cirrhosis, and AFP levels. Conclusion: Our data revealed that most of the NBNC PLC patients were ICC. Female patients with biliary inflammatory diseases and higher CA199 levels had an increased risk of ICC, and patients with metabolic risk factors and elevated AFP levels were more likely to develop HCC. Additional research should be performed to verify this finding.

[Dynamic changes of cellular HIV-1 DNA quantification during highly active antiretroviral therapy in Chinese HIV infected individuals].

OBJECTIVE: To observe the dynamic changes of 3 types of viral reservoir cells (NK cells, T lymphocytes and monocytes), and its relationship with treatment effect in Chinese HIV-1 infected patients receiving highly active antiretroviral treatment (HAART) for 2 years. METHODS: A total of 40 chronic HIV-1-infected adults who initiated HAART were enrolled in this study and followed up for 2 years. Peripheral whole blood was obtained from each patient at baseline (0 month), 6, 12, 18 and 24 months. Real-time fluorescent quantitative PCR was used to detect the HIV-1 RNA in the plasma and HIV-1 DNA in NK cells, T lymphocytes and monocytes. All the data were statistically analyzed. RESULTS: CD4 count increased with the decrease of the viral load during HAART. After HAART initiation, HIV-1 DNA showed a significant decrease in NK cells, T lymphocytes and monocytes. The HIV-1 DNA from T lymphocytes, NK cells and monocytes correlated positively with the HIV- 1 RNA (P<0.05) while NK cells and T lymphocytes correlated negatively with CD4+ T cell count. However we did not find significant correlation between CD4+ T cell count and HIV-1 DNA in monocytes at the baseline of HAART. CONCLUSION: This study found that NK cell was an important HIV cellular reservoir besides T lymphocytes and monocytes. T lymphocytes may be the main long lasting HIV reservoir. HIV-1 proviral DNA may play an important role in the efficacy of treatment and monitoring the disease progression.

Hepatitis B virus infection in patients with Wilson disease: A large retrospective study.

BACKGROUND: Wilson disease (WD) is the most common genetic metabolic liver disease. Some studies have shown that comorbidities may have important effects on WD. Data on hepatitis B virus (HBV) infection in patients with WD are limited. AIM: To investigate the prevalence and clinical impact of HBV infection in patients with WD. METHODS: The clinical data of patients with WD were analyzed retrospectively, and the data of patients with concurrent WD and HBV infection were compared with those of patients with isolated WD. RESULTS: Among a total of 915 WD patients recruited, the total prevalence of current and previous HBV infection was 2.1% [95% confidence interval (CI): 1.2%-3.0%] and 9.2% (95%CI: 7.3%-11.1%), respectively. The main finding of this study was the identification of 19 patients with concurrent WD and chronic hepatitis B (CHB) infection. The diagnosis of WD was missed in all but two patients with CHB infection. The mean delay in the diagnosis of WD in patients with concurrent WD and CHB infection was 32.5 mo, which was significantly longer than that in patients with isolated WD (10.5 mo). The rates of severe liver disease and mortality in patients with concurrent WD and CHB infection were significantly higher than those in patients with isolated WD (63.1% vs 19.3%, P = 0.000 and 36.8% vs 4.1%, P < 0.001, respectively). Binary logistic regression analysis revealed a significantly higher risk of severe liver disease at the diagnosis of WD in patients with current HBV infection [odds ratio (OR) = 7.748; 95%CI: 2.890-20.774; P = 0.000)] or previous HBV infection (OR = 5.525; 95%CI: 3.159-8.739; P = 0.000) than in patients with isolated WD. CONCLUSION: The total prevalence of current HBV infection in patients with WD was 2.1%. The diagnosis of WD in CHB patients is usually missed. HBV infection is an independent risk factor for severe liver disease in WD patients. The diagnosis of WD should be ruled out in some patients with CHB infection.

[Cellular HIV DNA quantitative testing and its significance in Chinese AIDS patients during 48 weeks' highly-active antiretroviral treatment].

OBJECTIVE: To observe the dynamic changes of peripheral blood T lymphocytes and monocytes, which serve as HIV-1 viral reservoirs, in Chinese HIV-infected patients receiving highly-active antiretroviral treatment (HAART) for 48 weeks and its clinical significance. METHODS: A total of 35 chronic HIV-1 infected adults initial received HAART. The peripheral blood T lymphocyte subsets counts were determined by flux cytometry at week 0, 24 and 48. Magnetic activated cell sorting was used to extract cellular DNA from monocytes and T lymphocytes purified from peripheral blood mononuclear cells. Real-time fluorescent quantitative PCR was used to detect the serum HIV RNA and HIV DNA of monocytes and T lymphocytes. SPSS 18.0 software was used to analyze the collected data. RESULTS: At week 0, 24, and 48 after initiation of HAART, HIV RNA levels of peripheral blood were (4.12 ± 1.41), ≤ 1.69, and ≤ 1.69 lg copies/ml, respectively; CD(4)(+) T cells were (196 ± 101), (321.90 ± 112) and (392 ± 127) cells/µl, respectively; HIV DNA level in T lymphocytes were (4.03 ± 0.53), (2.74 ± 1.16) and (2.45 ± 0.41) lg copies/10(6) cells respectively; while in monocytes, HIV DNA levels were (2.51 ± 0.68), (2.16 ± 0.34)and (2.03 ± 0.25)lg copies/10(6) cells. Statistical analysis revealed that HIV RNA level was negatively correlated with the CD(4)(+) T cell count through the whole trail, while positively correlated with the HIV DNA level in blood T lymphocytes and monocytes. HIV DNA level in T lymphocytes decreased more slowly than HIV DNA in monocytes. Moreover, peripheral blood CD(4)(+) T cell count was negatively associated with the HIV DNA capacity from T lymphocytes. CONCLUSIONS: Both T lymphocyte and monocyte may serve as viral reservoirs, and T lymphocyte might play a more important role as HIV reservoirs. The blood HIV RNA is correlated positively with the cellular HIV DNA, whereas, CD(4)(+) T cell count is correlated negatively with HIV DNA from lymphocytes, which suggests that HIV DNA levels in T lymphocyte might be one of indicators of AIDS progress during HAART.

Amphotericin B May Decrease the Serum Level of Voriconazole: A Case Report and Brief Review of Literature.

Background: Combination therapy with amphotericin B (AmB) and Voriconazole are sometimes used due to reducing AmB-related adverse reactions and improving outcomes. However, there was no report whether AmB would affect the serum level of Voriconazole. Case Presentation: A patient was in presumption of invasive fungal infection. Voriconazole and amphotericin B was combined for administration. The results showed that serum levels of Voriconazole were dramatically reduced when combined AmB, whereas went up when receiving Voriconazole alone. Conclusion: AmB might decrease the level of Voriconazole. Such a combination of AmB and Voriconazole cannot be considered appropriate until more data are available.

[Clinical characteristics of highly active antiretroviral therapy-associated immune reconstruction inflammation in acquired immunodeficiency syndrome].

OBJECTIVE: To summarize the morbidity, mortality, clinical manifestations and risk factors for IRIS (immune reconstruction inflammatory syndrome) during HAART (highly active antiretroviral therapy) in China. METHODS: From October 2007 to September 2009, a prospective cohort of 238 AIDS (acquired immunodeficiency syndrome) patients on HAART from Hunan and Jianxi provinces was recruited for a follow-up of 24 weeks. And 47 and 191 patients were assigned into the IRIS and non-IRIS groups respectively. The data of general information, clinical manifestations and treatment of two groups were collected and compared. Blood samples were collected in both groups at pre-and post-HAART 12 weeks, 24 weeks for HIV viral load and CD4(+) cell count examinations. A statistical analysis was performed. RESULTS: A total of 47 (19.7%) IRIS cases was analyzed. The median onset of IRIS was 28 (9 - 36) days. And 29 (61.7%) cases of tuberculosis IRIS were found. There was no significant difference in age, gender, route of transmission and antiretroviral regimens between the IRIS and non-IRIS groups. At baseline, Weeks 12 and 24, both groups showed a significant decline of viral load. And there was no significant difference between them. Both groups showed a significant increase of CD4(+) cell count. But there was no significant difference between two groups. However, the baseline CD4(+) cell count was markedly lower in the IRIS group than that in the non-IRIS group. In 85.1% (40/47) of cases, the CD4(+) cell count was < 100 × 10(6)/L in the IRIS group at the baseline of HAART. CONCLUSION: IRIS mostly occurs during 3 months of HAART initiation. The age, gender, route of transmission and antiretroviral treatment regimens of patients on HAART are not risk factors for the development of IRIS. The HIV RNA viral load decreases in both IRIS and non-IRIS groups without any significant difference. The patients with a CD4(+) cell count < 100/µl are more vulnerable to develop IRIS.

Prospect of IL-2, IL-7, IL-15 and IL-21 for HIV immune-based therapy.

Although highly active antiretroviral therapy (HAART) can effectively reduce the HIV replication, complete recovery of CD4(+) T cells does not always occur, even among patients with high virological control. Current researches on γ-chain cytokines have understood the biology and their crucial roles in initiating, maintaining, and regulating the immunologic homeostasis and the inflammatory processes. Due to the multiple functions such as the regulatory and effector cellular function in healthy and disease state, these molecules, their receptors, and their signal transduction pathways are promising candidates for therapeutic interference. The common γ-chain cytokines IL-2, IL-7, IL-15, and IL-21 are primary regulators of T cell homeostasis and thus have been considered prime immunotherapeutic candidates, both for increasing T cell levels/function and augmenting vaccine-elicited viral-specific T cell responses in immunocompromised AIDS patients. The Objective of this review is to update the role of the common γ-chain cytokines IL-2, IL-7, IL-15, and IL-21 in HIV AIDS pathogenesis.

IL-21 level in Chinese HIV infected individuals and its dynamics undergoing HAART.

OBJECTIVE: To investigate the dynamics of interleukin-21(IL-21) cytokine in the Chinese HIV patients undergoing highly active antiretroviral therapy (HAAPT). METHODS: A total of 25 adults with chronic HIV infections, responding to combined highly active antiretroviral therapy (HAART) guideline criteria were enrolled for a 1-year follow-up. After signing an informed consent, 20 mL blood was collected from each patient at the base line, 6 month and 12 month, respectively. CD4 and CD8 cell count was quantified by flux cytometry, serum HIV RNA quantified by real time PCR and IL-21 concentrations by ELISA. RESULTS: IL-21 levels increased gradually during the follow-up but did not reach the healthy levels. IL-21 correlated positively with the CD4 cells but not with CD8 T cells. HIV RNA correlated negatively with CD4 cell count but did not show any relationship with the CD8 cells. CONCLUSION: IL-21 has potential role in the immunopathogenesis of HIV, and might be an important factor in immune construction during HAART.

Evaluation of a 6-year highly active antiretroviral therapy in Chinese HIV-1-infected patients.

OBJECTIVE: To evaluate the long-term efficacy and tolerability of nevirapine (NVP)-based regimens in the treatment of human immunodeficiency virus (HIV)-infected Chinese patients in routine clinical practice. METHODS: From October 2002 to May 2004, 57 HIV-1-infected patients commenced antiretroviral therapy (ART), and were followed up to December 2008. These antiretroviral-naïve patients, who originally received two nucleoside reverse transcriptase inhibitors and NVP, had HIV RNA levels, T lymphocyte subsets and safety parameters assessed over 6 years. RESULTS: Of the 57 patients, 34 patients participated in the long-term follow-up. After 5-6 years, >60% of the patients had HIV RNA levels <50 copies/microl, and the median increase in CD4 cell counts from baseline was 329 cells/microl. gamma-Glutamyl transferase increased in 17 patients (29.8%); serum cholesterol and triglyceride levels were elevated in 15 patients (26.3%), and 25.0% (6/24) of the patients developed lipodystrophy (mainly females). Grade 3/4 adverse events occurred in 3 cases. CONCLUSION: ART with NVP-based regimens suppressed HIV viremia and produced continued CD4 cell increases in a majority of subjects for 6 years. Safety and tolerance were good with no unexpected long-term toxicity. Though based on a small group, this study demonstrates durable effects of ART in Chinese patients.

[Effects of different mutated sites in vpr gene of HIV on apoptosis of host cells: experiment with HeLa cells].

OBJECTIVE: To investigate the effects of different mutated sites in the vpr gene of HIV on the apoptosis of host cells, and the possible mechanism thereof. METHODS: Fourteen HIV-1 vpr fragments were obtained from HIV-infected persons. Eukaryotic expression vector pcDNA3.1 (+) plasmid was extracted, the PCR purified product was double-cut by HindIII and BamH, and the cut products were ligated to vectors, thus establishing the JM109 competent cells. Sequencing was used to confirm the reconstruction of pcDNA-vpr eukaryotic expression vectors that were then transfected into HeLa cells. Blank vectors were transfected as control group. Cells were harvested after 24 hours and underwent Hoechst 33258 staining and observed under fluorescence microscope. Annexin-FITC-PI staining and flow cytometry were used to observe the percentage of apoptosis. The caspase-3 activity was detected by enzyme labeling instrument. RESULTS: The apoptotic rates shown by Hoechst and annexin--FITC-PI staining methods, and caspase-3 activity levels of the HeLa cells transfected with the gene fragments with mutated sites 70, 85, 86, and 94 cells were all lower than the cells transfected with the gene fragments without these mutated sites. The apoptosis causing ability levels of the No 1-7 recombinant plasmids (all of the Vpr AE subtype) were all lower than those of the No 8-14 plasmids (of Vpr B, AB, C, and C/BC subtypes). CONCLUSION: The apoptosis causing ability of the HIV with the vpr sequence with mutated sites 70, 85, 86, 94 is significantly lower than those without these sites. AE subtype induces lower apoptotic behavior in the hoist cells, and decreased activation of the caspase-3 pathway may be one of the mechanisms.

[Therapeutic effect and safety evaluation on 6-year highly active antiretroviral therapy for Chinese HIV-1 infected patients].

OBJECTIVE: To evaluate the long-term efficacy and safety of nevirapine (NVP)-based regimens for HIV-infected Chinese patients in routine clinical practice. METHODS: From October 2002 to May 2004, 57 HIV-1-infected patients commenced highly active antiretroviral therapy (HAART), and were followed to December 2008. They originally received 2 nucleoside reverse transcriptase inhibitors (NRTIs) and nevirapine. HIV RNA levels, T lymphocyte subsets and safety were assessed. Blood routine test and main laboratory parameter changes were traced. If apparent side effects or virological failure appeared we would, if necessary, terminate the therapy or change the regimen. RESULTS: Of the 57 subjects, 34 were followed-up for more than 4 years. After 5-6 years, 63.3% of the subjects (19/30) had HIV RNA levels<50 copies/microL, and the median increase in CD4(+) cell count from the baseline was 329 cells/microL. The mean decrease in CD8(+) cell count was 128 cells/microL. At the same time, the CD4(+) CD45RA+CD62L cell count and CD4(+)CD45RO(+) cell gradually increased, and the counts of CD8(+)CD38(+) cell declined gradually. These changes are apparent 2 years after HAART. The increase rate slowed down after 2 years. But they did not recover completely as well as healthy people at year 6. About 56% (32/57) of HIV-infected patients developed various drug-related side effects. The most common was gastrointestinal side effect, followed nervous disorder, baldness, and rashes, mostly happened in 6 months. Gamma-GT increased occurred in 29.8% of patients (17/57), and serum cholesterol and triglyceride elevated in 26.3% of the patients (15/57). Six patients developed lipodystrophy, mainly in female patients, and 25 patients showed abnormal blood picture and liver function, renal function changes and amylase elevation. Grade 3-4 adverse events occurred in 3 cases (2 peripheral neuropathy, and 1 suspected lactic acidosis). One subject experienced grade 3 rashes. CONCLUSION: Antiretroviral therapy with NVP-based regimens is safe and effective by suppressing HIV viremia and producing continued CD4 cell increases in subjects with HIV or AIDS for 6 years.

Identifying factors affecting the pharmacokinetics of voriconazole in patients with liver dysfunction: A population pharmacokinetic approach.

Voriconazole is a broad-spectrum antifungal agent commonly used to treat invasive fungal infections. Voriconazole has significant intraindividual and interindividual pharmacokinetics variability in different patient populations. Pharmacokinetic data of voriconazole in patients with liver dysfunction were limited. The aims of this study were to evaluate the population pharmacokinetics of voriconazole in patients with liver dysfunction and to identify the factors that affect voriconazole pharmacokinetics. A total of 166 samples taken from 57 patients with liver dysfunction were included in the study. A one-compartment pharmacokinetic model with first-order absorption and elimination was used to describe the data. Voriconazole clearance (CL) was 0.58 L/h, the volume of distribution (Vd ) was 134 L, and oral bioavailability (F) was 80.8%. This study showed that platelet count was significantly associated with voriconazole pharmacokinetic parameters. CYP2C19 polymorphisms had no effect on voriconazole pharmacokinetic parameters. Voriconazole CL was significantly decreased in patients with liver dysfunction. This study provides useful pharmacokinetics information for patients with liver dysfunction while highlighting the value of therapeutic drug monitoring in adjusting doses.

Cryptococcosis manifesting as isolated biliary infection: Case report and brief review of literature.

BACKGROUND: Biliary cryptococci infection is rare, which is frequently diagnosed by exploratory laparotomy, preoperative diagnosis is difficult. CASE PRESENTATION: A 14-year-old girl presented with intermittent jaundice for 6 years. She had no pruritus, anorexia, nausea or vomiting, fever, abdominal pain, or clay stools. Laboratory tests showed obstructive jaundice, eosinophilia, and increased IgE levels. The patient was ultimately diagnosed as Cryptococcal infection by bone marrow culture. The patient responded to antifungal therapy. CONCLUSION: Unnecessary surgical intervention was avoided by an early and accurate diagnosis. Cryptococcosis infection of bile duct should be highly suspected, when the children with obstructive jaundice have eosinophilia and increased IgE levels.

Evaluation of human immunodeficiency virus type 1-infected Chinese patients treated with highly active antiretroviral therapy for two years.

The purpose of this study was to evaluate the long-term efficacy and safety of nevirapine in combination with didanosine and stavudine in the treatment of human immunodeficiency virus (HIV)1-infected Chinese patients in routine clinical practice. The study, from April 2003 to May 2005, with follow-up through 24 mo, was conducted at the Department of Infectious Diseases, Second Xiangya Hospital, Central-South University in Changsha, Hunan Province, China. Twenty-seven HIV1-infected patients received didanosine, stavudine, and nevirapine. Information from case notes regarding age, sex, side effects, viral load, naive and memory T cells, and CD4(+) and CD8(+) T cell count at baseline, 3, 6, 12, 18, and 24 mo was collected and analyzed. Virologic suppression, defined as an HIV RNA concentration of less than 50 copies/mL at months 3, 6, 12, 18, and 24, was considered the main outcome measure. Of 27 patients, 17 were men with a mean age 33.5 yr. The mean baseline viral load was 5.15 log copies/mL and the mean CD4(+) cell count was 185 cells/dL. Of 27 patients, 3 patients discontinued study medication; treatment was changed, because of side effects, from didanosine (ddI), stavudine (d4T), and nevirapine (NVP) to zidovudine, lamivudine, and NVP for 24 patients who had completed 24 mo of treatment with ddI, d4T, and NVP; and viral load suppression was attained in 17 patients (70.8%) at 12 mo, in 14 patients (58.3%) at 18 mo, and in 13 patients (56.6%) at 24 mo. The CD4 T cell count increased by 114 cells/microL (mean, 299 cells/microL) after 12 mo of treatment and by 132 cells/microL (mean, 317 cells/microL) after 24 mo of treatment. Naive T cells and memory cells also increased in number, but at a slower rate. Activated (CD38(+)) CD8(+) T cells were elevated at baseline (67.7%) and declined by month 24 (49.7%), but did not reach normal levels. We conclude that a regimen of NVP with ddI and d4T provided durable suppression of plasma viral load in HIV-infected patients, with significant improvement in the CD4 cell count, and can be well tolerated by patients with HIV-1 infection.

Role of autophagy in tumor necrosis factor-α-induced apoptosis of osteoblast cells.

The aim of this study is to investigate the role of tumor necrosis factor-α (TNF-α) in apoptosis and autophagy of mouse osteoblast MC3T3-E1 cells, as well as the crosstalk between autophagy and apoptosis. Mouse osteoblast MC3T3-E1 cells were cultured in vitro and treated with 5-fluorouracil (5-FU), rapamycin, 3-methyl adenine (3-MA) and TNF-α either alone or in combination, respectively. MTT assays were used to monitor the cell viability upon different treatments. Annexin-V-FITC/propidium iodide (PI) staining was used to detect the apoptotic rate of osteoblasts. Autophagic structure and apoptotic bodies were visualized by transmission electron microscopy (TEM). Western blot analysis was performed to detect the autophagic marker LC3-II/I, p62 and apoptotic marker cleaved caspase-3. TNF-α inhibits MC3T3-E1 cell viability in a dose-dependent and time-dependent manner. Annexin-V-FITC/PI staining, coupled with TEM, showed that TNF-α induced cell apoptosis and autophagy in MC3T3-E1 cells. The autophagy inducer rapamycin ameliorated TNF-α-induced apoptosis. In contrast, 3-MA, which is an autophagy inhibitor, caused an exaggerated induction of TNF-α-induced apoptosis. TNF-α upregulated autophagy marker LC3-II/I, but downregulated p62 in osteoblasts. Combined treatment of rapamycin and TNF-α further exaggerated this effect, whereas co-treatment of 3-MA and TNF-α decreased LC3-II/I, but increased p62 compared with TNF-α alone. In addition, TNF-α caused an induction of apoptotic marker cleaved caspase-3. TNF-α-mediated induction of cleaved caspase-3 was downregulated by rapamycin, but upregulated by 3-MA, respectively. TNF-α induced both autophagy and apoptosis in osteoblasts, and upregulated autophagy protects the cell by reducing TNF-α-induced apoptosis.