Mitochondrial Bioenergetic and Proteomic Phenotyping Reveals Organ-Specific Consequences of Chronic Kidney Disease in Mice.

Chronic kidney disease (CKD) results in reduced kidney function, uremia, and accumulation of uremic metabolites. Mitochondrial alterations have been suggested to play a role in the disease pathology within various tissues. The purpose of this study was to perform a comprehensive bioenergetic and proteomic phenotyping of mitochondria from skeletal muscle (SkM), cardiac muscle (CM), and renal tissue from mice with CKD. The 5-month-old C57BL/6J male mice were fed a casein control or adenine-supplemented diet for 6 months. CKD was confirmed by blood urea nitrogen. A mitochondrial diagnostic workflow was employed to examine respiratory function, membrane and redox potential, reactive oxygen species production, and maximal activities of matrix dehydrogenases and electron transport system (ETS) protein complexes. Additionally, tandem-mass-tag-assisted proteomic analyses were performed to uncover possible differences in mitochondrial protein abundance. CKD negatively impacted mitochondrial energy transduction (all p < 0.05) in SkM, CM, and renal mitochondria, when assessed at physiologically relevant cellular energy demands (ΔGATP) and revealed the tissue-specific impact of CKD on mitochondrial health. Proteomic analyses indicated significant abundance changes in CM and renal mitochondria (115 and 164 proteins, p < 0.05), but no differences in SkM. Taken together, these findings reveal the tissue-specific impact of chronic renal insufficiency on mitochondrial health.

Inhibition of microbiota-dependent TMAO production attenuates chronic kidney disease in mice.

Patients with chronic kidney disease (CKD) have elevated circulating levels of trimethylamine N-oxide (TMAO), a metabolite derived from gut microbes and associated with cardiovascular diseases. High circulating levels of TMAO and its dietary precursor, choline, predict increased risk for development of CKD in apparently healthy subjects, and studies in mice fed TMAO or choline suggest that TMAO can contribute to kidney impairment and renal fibrosis. Here we examined the interactions between TMAO, kidney disease, and cardiovascular disease in mouse models. We observed that while female hyperlipidemic apoE KO mice fed a 0.2% adenine diet for 14 weeks developed CKD with elevated plasma levels of TMAO, provision of a non-lethal inhibitor of gut microbial trimethylamine (TMA) production, iodomethylcholine (IMC), significantly reduced multiple markers of renal injury (plasma creatinine, cystatin C, FGF23, and TMAO), reduced histopathologic evidence of fibrosis, and markedly attenuated development of microalbuminuria. In addition, while the adenine-induced CKD model significantly increased heart weight, a surrogate marker for myocardial hypertrophy, this was largely prevented by IMC supplementation. Surprisingly, adenine feeding did not increase atherosclerosis and significantly decreased the expression of inflammatory genes in the aorta compared to the control groups, effects unrelated to TMAO levels. Our data demonstrate that inhibition of TMAO production attenuated CKD development and cardiac hypertrophy in mice, suggesting that TMAO reduction may be a novel strategy in treating CKD and its cardiovascular disease complications.

Alteration of serum amino acid profiles by dietary adenine supplementation inhibits fatty liver development in rats.

Studies on animal models have demonstrated that feeding a low-arginine diet inhibits triacylglycerol (TAG) secretion from the liver, resulting in marked fatty liver development in rats. Here, we first showed that culturing hepatocytes in the medium mimicking the serum amino acid profile of low-arginine diet-fed rats induced TAG accumulation in the cells, indicating that the specific amino acid profile caused TAG accumulation in hepatocytes. Dietary adenine supplementation completely recovered hepatic TAG secretion and abolished hepatic TAG accumulation in rats. A comprehensive non-linear analysis revealed that inhibition of hepatic TAG accumulation by dietary adenine supplementation could be predicted using only serum amino acid concentration data. Comparison of serum amino acid concentrations indicated that histidine, methionine, and branched-chain amino acid (BCAA) concentrations were altered by adenine supplementation. Furthermore, when the serum amino acid profiles of low-arginine diet-fed rats were altered by modifying methionine or BCAA concentrations in their diets, their hepatic TAG accumulation was abolished. Altogether, these results suggest that an increase in methionine and BCAA levels in the serum in response to dietary arginine deficiency is a key causative factor for hepatic TAG accumulation, and dietary adenine supplementation could disrupt this phenomenon by altering serum amino acid profiles.

Efficacy of high-dose corticosteroid-based treatment for chronic lymphocytic leukemia patients with p53 abnormalities in the era of B-cell receptor inhibitors.

PURPOSE: High-dose methylprednisolone (HDMP) with or without anti-CD20 antibody treatment in the pre B-cell receptor inhibitor (BCRi) era was used as potential salvage therapy for relapsed/refractory chronic lymphocytic leukemia/small lymphocytic lymphoma (r/r CLL/SLL) patients bearing the 17p deletion. PATIENTS AND METHODS: Outcomes were compared in retrospect between r/r patients treated with HDMP (n = 20), ibrutinib (n = 39) and idelalisib with rituximab (n = 14). RESULTS: Higher overall response rates were found in those patients undergoing BCRi therapy compared to HDMP (79.2% vs. 0%; p < 0.0001), along with longer median progression-free survival (not reached vs. 24.1 months; p < 0.01). Nevertheless, there were no differences in the overall survival (HDMP 35.87 months vs. not reached; p = 0.58). CONCLUSION: HDMP treatment was significantly inferior in terms of response rate and progression-free survival in r/r CLL/SLL patients with the 17p deletion, and may only be used whenever novel compounds are unavailable.

Pleural fluid MYD88 L265P mutation supporting diagnosis and decision to treat extramedullary Waldenstrom's macroglobulinemia: a case report.

BACKGROUND: Our case of a patient with untreated lymphoplasmacytic lymphoma/Waldenstrom's macroglobulinemia with extramedullary pleural effusion is the first documented case of pleural fluid MYD88 L265P mutation status in a community hospital setting. Our patient was intolerant to 420 mg ibrutinib, but still achieved a lasting complete remission, as defined by National Comprehensive Cancer Network guidelines, with a dose reduction to 240 mg of ibrutinib. CASE PRESENTATION: A 72-year-old Caucasian (white) man diagnosed with monoclonal immunoglobin M kappa lymphoplasmacytic lymphoma/Waldenstrom's macroglobulinemia monitored without treatment for 2 years, presented with dyspnea and a left pleural effusion. At presentation, computed tomography scans of his chest, abdomen, and pelvis showed layering left pleural effusion and para-aortic lymphadenopathy. Pleural fluid cytology demonstrated B-cell lymphoma of the lymphoplasmacytic subtype, with monoclonal kappa B-cell population on flow and a positive MYD88 L265P mutation. The pleural effusion recurred post-thoracentesis and he achieved a lasting complete remission as defined by National Comprehensive Cancer Network guideline with 240 mg ibrutinib. CONCLUSIONS: Our discussion details a comprehensive literature review of extramedullary pulmonary involvement in Waldenstrom's macroglobulinemia. Establishing a malignant etiology for pleural effusion in Waldenstrom's macroglobulinemia can be challenging, as standard techniques may be insensitive. Allele-specific polymerase chain reaction for detecting MYD88 L265P mutations is more sensitive for confirming lymphoplasmacytic lymphoma/Waldenstrom's macroglobulinemia in pleural fluid. Extramedullary pulmonary involvement usually presents post-diagnosis of Waldenstrom's macroglobulinemia and responds well to Waldenstrom's macroglobulinemia-directed treatment regimens. Allele-specific polymerase chain reaction is a sensitive assay for detecting MYD88 L265P mutations in pleural fluid to support the diagnosis of malignant pleural effusion in the setting of Waldenstrom's macroglobulinemia and helps guide the treatment decision to use ibrutinib. Although intolerant of ibrutinib 420 mg, our patient achieved complete and sustained remission of pleural effusion with a dose of 240 mg with progression free survival of over 30 months.

Self-assembled nucleo-tripeptide hydrogels provide local and sustained doxorubicin release.

Self-assembled nucleo-peptide hydrogels have a nanofibril structure composed of noncovalent molecular interactions between peptide groups as well as π-π stacking and Watson-Crick interactions via complementary nucleobases. These hydrogels have specific benefits for biomedical applications due to their DNA-like interactions in addition to the well-known advantages of peptide biomaterials: biocompatibility, extracellular matrix (ECM)-like structure, and bottom-up design. Inspired by the nucleobase stacking structure, we hypothesized that nucleo-peptides would be able to deliver the DNA-intercalating chemotherapeutic, doxorubicin (Dox) in a sustained manner when delivered locally to a solid tumor. Ade-FFF nucleo-peptide hydrogels were able to load a high concentration of Dox (1 mM) and demonstrated continuous release under in vitro degradation conditions. We adopted an in vivo tumor-bearing mouse model to evaluate the delivery of Dox by Ade-FFF hydrogels. We found that Dox-containing hydrogels reduced tumor growth and resulted in greater apoptosis-mediated cell death in the tumor as evidenced by caspase-3 expression. Pharmacokinetics and biodistribution studies also supported the observation that Dox delivery by an Ade-FFF hydrogel improves sustained delivery in the local tumor site. This study demonstrates the potential of self-assembled nucleo-peptides in biomedical applications by using their distinctive DNA-like structure.

Successful use of once-daily high-dose darunavir and dolutegravir in multidrug-resistant HIV.

WHAT IS KNOWN AND OBJECTIVE?: Antiretroviral (ARV) resistance may result during periods of consistently poor adherence. We report the successful use of a novel once-daily (QD) ARV regimen in a patient with multidrug-resistant (MDR) HIV. CASE SUMMARY: Once-daily darunavir 1200 mg/ritonavir 100 mg, dolutegravir and emtricitabine/tenofovir alafenamide was initiated with directly observed therapy. With the assistance of therapeutic drug monitoring, dolutegravir dosing was increased to 150 mg daily. The patient maintained virologic suppression for 18 months. WHAT IS NEW AND CONCLUSIONS?: In this case, QD darunavir/ritonavir achieved similar trough concentrations to twice daily dosing with dolutegravir dose titration necessitated and resulted in HIV virologic control.

Chronic treatment of curcumin improves hepatic lipid metabolism and alleviates the renal damage in adenine-induced chronic kidney disease in Sprague-Dawley rats.

BACKGROUND: Chronic kidney disease (CKD), including nephrotic syndrome, is a major cause of cardiovascular morbidity and mortality. The literature indicates that CKD is associated with profound lipid disorders due to the dysregulation of lipoprotein metabolism which progresses kidney disease. The objective of this study is to evaluate the protective effects of curcumin on dyslipidaemia associated with adenine-induced chronic kidney disease in rats. METHODS: Male SD rats (n = 29) were divided into 5 groups for 24 days: normal control (n = 5, normal diet), CKD control (n = 6, 0.75% w/w adenine-supplemented diet), CUR 50 (n = 6, 50 mg/kg/day curcumin + 0.75% w/w adenine-supplemented diet), CUR 100 (n = 6, 100 mg/kg/day curcumin + 0.75% w/w adenine-supplemented diet), and CUR 150 (n = 6, 150 mg/kg/day curcumin + 0.75% w/w adenine-supplemented diet). The serum and tissue lipid profile, as well as the kidney function test, were measured using commercial diagnostic kits. RESULTS: The marked rise in total cholesterol, low-density lipoprotein (LDL) cholesterol, very low-density lipoprotein (VLDL) cholesterol, triglycerides and free fatty acids in serum, as well as hepatic cholesterol, triglyceride and free fatty acids of CKD control rats were significantly protected by curcumin co-treatment (at the dose of 50, 100 and 150 mg/kg). Furthermore, curcumin significantly increased the serum high-density lipoprotein (HDL) cholesterol compared to the CKD control rats but did not attenuate the CKD-induced weight retardation. Mathematical computational analysis revealed that curcumin significantly reduced indicators for the risk of atherosclerotic lesions (atherogenic index) and coronary atherogenesis (coronary risk index). In addition, curcumin improved kidney function as shown by the reduction in proteinuria and improvement in creatinine clearance. CONCLUSION: The results provide new scientific evidence for the use of curcumin in CKD-associated dyslipidaemia and substantiates the traditional use of curcumin in preventing kidney damage.

Chronic kidney disease induced by an adenine rich diet upregulates integrin linked kinase (ILK) and its depletion prevents the disease progression.

Kidney fibrosis is one of the main pathological findings of progressive chronic kidney disease (CKD) although the pathogenesis of renal scar formation remains incompletely explained. Integrin-linked kinase (ILK), a major scaffold protein between the extracellular matrix (ECM) and intracellular signaling pathways, is involved in several pathophysiological processes during renal damage. However, ILK contribution in the CKD progress remains to be fully elucidated. In the present work, we studied 1) the renal functional and structural consequences of CKD genesis and progression when ILK is depleted and 2) the potential of ILK depletion as a therapeutic approach to delay CKD progression. We induced an experimental CKD model, based on an adenine-supplemented diet on adult wild-type (WT) and ILK-depleted mice, with a tubulointerstitial damage profile resembling that is observed in human CKD. The adenine diet induced in WT mice a progressive increase in plasma creatinine and urea concentrations. In the renal cortex it was also observed tubular damage, interstitial fibrosis and progressive increased ECM components, pro-inflammatory and chemo-attractant cytokines, EMT markers and TGF-ß1 expressions. These observations were highly correlated to a simultaneous increase of ILK expression and activity. In adenine-fed transgenic ILK-depleted mice, all these changes were prevented. Additionally, we evaluated the potential role of ILK depletion to be applied after the disease induction, as an effective approach to interventions in human CKD subjects. In this scenario, two weeks after the establishment of adenine-induced CKD, ILK was abrogated in WT mice and stabilized renal damage, avoiding CKD progression. We propose ILK to be a potential target to delay renal disease progression.

Telbivudine and adefovir dipivoxil combination therapy improves renal function in patients with chronic hepatitis B: A STROBE-compliant article.

Few studies have addressed the impact of adefovir dipivoxil (ADV)-based combination therapy on the renal function of patients with chronic hepatitis B (CHB). This study evaluated the effects of ADV combined with other antiviral nucleotide analogs (NAs) on renal function of patients with CHB, and analyzed the risk factors for more than 20% reduction of baseline estimated glomerular filtration rate (eGFR).The data of 164 patients with CHB were retrospectively analyzed in this study. Of the 164 patients, 42 received entecavir (ETV) combined with ADV (ETV + ADV group), 68 lamivudine (LAM) combined with ADV (LAM + ADV group), and 54 telbivudine (LDT) combined with ADV (LDT + ADV group). Serum creatinine (SCr) level, eGFR, and proportion of patients with eGFR ≥ 90 mL/min/1.73 m were observed. Also, the independent risk factors for more than 20% reduction of baseline eGFR were analyzed.After 104-week combination therapy, compared with the baseline level, SCr levels were significantly increased in the ETV + ADV group (67 µmol/L vs 73 µmol/L, P = .012) and LAM + ADV group (68 µmol/L vs 79 µmol/L, P = .008), but it was significantly decreased in the LDT + ADV group (69 µmol/L vs 64 µmol/L, P = .023). Compared with the baseline level, eGFR was significantly decreased in the ETV + ADV group (107.8 mL/min/1.73 m vs 96.1 mL/min 1.73/m, P = .004), and LAM + ADV group (105.4 mL/min/1.73 m vs 87.3 mL/min/1.73 m, P = .000), but it was significantly increased in the LDT + ADV group (104.1 mL/min 1.73/m vs 116.2 mL/min/1.73 m,P = .005). The proportion of patients with normal renal function (≥90 mL/min/1.73 m) was significantly higher in the LDT + ADV group than in the ETV + ADV group (P = .002) and LAM + ADV group (P = .001). Multivariate analysis showed that age (P = .035), male (P = .005), baseline eGFR (P < .001), LAM combined with ADV (P < .008), and ETV combined with ADV (P = .03) were independent risk factors for 20% reduction of baseline eGFR.As compared with ETV and ADV combination therapy as well as LAM and ADV combination therapy, LDT and ADV combination therapy can improve eGFR level, so LDT and ADV combination therapy is suitable for the patients with potential renal impairment.

Human non­small cell lung cancer cells can be sensitized to camptothecin by modulating autophagy.

Lung cancer is a prevalent disease and is one of the leading causes of mortality worldwide. Despite the development of various anticancer drugs, the prognosis of lung cancer is relatively poor. Metastasis of lung cancer, as well as chemoresistance, is associated with a high mortality rate for patients with lung cancer. Camptothecin (CPT) is a well-known anticancer drug, which causes cancer cell apoptosis via the induction of DNA damage; however, the cytotoxicity of CPT easily reaches a plateau at a relatively high dose in lung cancer cells, thus limiting its efficacy. The present study demonstrated that CPT may induce autophagy in two human non­small cell lung cancer cell lines, H1299 and H460. In addition, the results of a viability assay and Annexin V staining revealed that CPT-induced autophagy could protect lung cancer cells from programmed cell death. Conversely, the cytotoxicity of CPT was increased when autophagy was blocked by 3-methyladenine treatment. Furthermore, inhibition of autophagy enhanced the levels of CPT-induced DNA damage in the lung cancer cell lines. Accordingly, these findings suggested that autophagy exerts a protective role in CPT-treated lung cancer cells, and the combination of CPT with a specific inhibitor of autophagy may be considered a promising strategy for the future treatment of lung cancer.

Serial Culture Is Critical for In Vitro Development of Parthenogenetic Embryos in the Golden Syrian Hamster.

Unlike oocytes of many other mammalian species, parthenogenetically activated hamster oocytes have not been reported to develop beyond the two-cell stage. This study investigated the in vitro development into blastocysts of parthenogenetic embryos of Golden Syrian hamsters. We observed that hamster oocytes could easily be artificially activated (AA) by treatment with ionomycin plus 6-dimethylaminopurine + cycloheximide + cytochalasin B as assessed by embryo cleavage in HECM-9 (63.15%) or HECM-10 (63.82%). None of the cleaved embryos developed beyond the two-cell stage when cultured in either of the two media. However, some of the embryos overcame the two-cell block and developed to the blastocyst stage (26.45%) when they were first cultured in HECM-10 for 24 hours and then in HECM-9 (serial culture media HECM-10-9) for 72 hours. Blastocyst development was further significantly (66.2%) improved when embryos were cultured in HECM-10 supplemented with ethylenediaminetetraacetic acid for 24 hours, then in HECM-9 supplemented with glucose for 72 hours (serial culture media HECM-11a-b). Hamster oocytes activated with ionomycin, ethanol, or a combination of the two treatments would develop to the blastocyst stage in serial culture media HECM-11a-b, whereas none of the spontaneously activated oocytes cleaved (0% vs. 86.93%, p < 0.05). DNA and microtubule configurations of spontaneously activated and AA oocytes were assessed by immunocytochemical staining and fluorescence microscopy. The results indicate that serial culture and the method of activation are critical for overcoming the in vitro developmental block of hamster parthenogenetic embryos. This study is the first to report blastocyst development from parthenogenetically activated hamster oocytes.

USC-087 protects Syrian hamsters against lethal challenge with human species C adenoviruses.

Human adenoviruses (AdV) cause generally mild infections of the respiratory and GI tracts as well as some other tissues. However, AdV can cause serious infection in severely immunosuppressed individuals, especially pediatric patients undergoing allogeneic hematopoietic stem cell transplantation, where mortality rates are up to 80% with disseminated disease. Despite the seriousness of AdV disease, there are no drugs approved specifically to treat AdV infections. We report here that USC-087, an N-alkyl tyrosinamide phosphonate ester prodrug of HPMPA, the adenine analog of cidofovir, is highly effective against multiple AdV types in cell culture. USC-087 is also effective against AdV-C6 in our immunosuppressed permissive Syrian hamster model. In this model, hamsters are immunosuppressed by treatment with high dose cyclophosphamide. Injection of AdV-C6 (or AdV-C5) intravenously leads to a disseminated infection that resembles the disease seen in humans, including death. We have tested the efficacy of orally-administered USC-087 against the median lethal dose of intravenously administered AdV-C6. USC-087 completely prevented or significantly decreased mortality when administered up to 4 days post challenge. USC-087 also prevented or significantly decreased liver damage caused by AdV-C6 infection, and suppressed virus replication even when administered 4 days post challenge. These results imply that USC-087 is a promising candidate for drug development against HAdV infections.

Triggering of Autophagy by Baicalein in Response to Apoptosis after Spinal Cord Injury: Possible Involvement of the PI3K Activation.

High level apoptosis induced by spinal cord injury (SCI) evokes serious damage because of the loss and dysfunction of motor neurons. Our previous studies showed that inhibition of autophagy evokes the activation of apoptosis. Interestingly, Baicalein, a medicine with anti-apoptosis activity that is derived from the roots of herb Scutellaria baicalensis, largely induces autophagy by activating phosphatidylinositol 3-kinase (PI3K). In this study, we investigated the effects of intraperitoneal injection of Baicalein on autophagy and apoptosis in SCI mice and evaluated the relationship between autophagy and apoptosis. We demonstrated that Baicalein promoted the functional recovery of motor neurons at 7 d after SCI. In addition, Baicalein enhanced neuronal autophagy and the autophagy-related factor PI3K, while inhibiting the p62 protein. Baicalein treatment decreased neuronal apoptosis at 7 d after SCI. Moreover, when inhibiting autophagy, apoptosis was upgraded by Baicalein treatment after injury. Thus, Baicalein attenuated SCI by inducing autophagy to reduce apoptosis in neurons potentially via activating PI3K.

Clinical features of hypophosphatemic osteomalacia induced by long-term low-dose adefovir dipivoxil.

OBJECTIVE: To investigate the predictors of hypophosphatemic osteomalacia induced by adefovir dipivoxil (ADV) and to monitor for early detection. PATIENTS AND METHODS: Hospitalized patients who were diagnosed with ADV-related hypo-phosphatemic osteomalacia were recruited and retrospectively analyzed in our hospital from January 2012 to December 2016. A telephone interview was conducted at 1, 3, 6, 9, 12, and 24 months after cessation of ADV. RESULTS: In the 8 patients enrolled in the study, the hypophosphatemic osteomalacia symptoms developed at an average of 5.14 (4-7) years since ADV treatment (10 mg/d). The average alkaline phosphatase (ALP) level was 279.50 (137-548) U/L, which was significantly higher than the normal level (45-125 U/L). The serum phosphorus level was an average of 0.59 (0.43-0.69) mmol/L, which was lower than the normal range (2.06-2.60 mmol/L). Serum calcium levels of the enrolled patients remained within normal limits. Reduced estimated glomerular filtration rate (eGFR <29 mL/min/1.73 m2) was seen in 4 cases. The clinical manifestations were mainly progressive systemic bone and joint pain, frequent fractures, trouble in walking, height reduction (4-6 cm), and so on. After cessation of ADV, symptoms like bone pain resolved gradually. Serum phosphorus level restored to normal in 4.5 months after the withdrawal of ADV. However, in 4 patients, renal function failed to return to normal in 24 months. CONCLUSION: More attention should be paid to the duration of ADV treatment. The level of serum phosphorus and ALP, as well as renal function, should be monitored for early detection of potential adverse drug reactions.

Hypo-osmolar Formulation of Tenofovir (TFV) Enema Promotes Uptake and Metabolism of TFV in Tissues, Leading to Prevention of SHIV/SIV Infection.

Oral preexposure prophylaxis (PrEP) has been approved for prophylaxis of HIV-1 transmission but is associated with high costs and issues of adherence. Protection from anal transmission of HIV using topical microbicides and methods congruent with sexual behavior offers the promise of improved adherence. We compared the pharmacokinetics (PK) and ex vivo efficacy of iso-osmolar (IOsm) and hypo-osmolar (HOsm) rectal enema formulations of tenofovir (TFV) in rhesus macaques. Single-dose PK of IOsm or HOsm high-dose (5.28 mg/ml) and low-dose (1.76 mg/ml) formulations of TFV enemas were evaluated for systemic uptake in blood, colorectal biopsy specimens, and rectal CD4+ T cells. Markedly higher TFV concentrations were observed in plasma and tissues after administration of the HOsm high-dose formulation than with all other formulations tested. TFV and TFV diphosphate (TFV-DP) concentrations in tissue correlated for the HOsm high-dose formulation, demonstrating rapid uptake and transformation of TFV to TFV-DP in tissues. TFV-DP amounts in tissues collected at 1 and 24 h were 7 times and 5 times higher, respectively (P < 0.01), than the ones collected in tissues with the IOsm formulation. The HOsm high-dose formulation prevented infection in ex vivo challenges of rectal tissues collected at 1, 24, and 72 h after the intrarectal dosing, whereas the same TFV dose formulated as an IOsm enema was less effective.

Cost-effectiveness analysis of tenofovir disoproxil fumarate for treatment of chronic hepatitis B in China.

BACKGROUND: Tenofovir disoproxil fumarate (TDF) is newly available for treatment of chronic hepatitis B patients in China. To date, no study has been conducted to examine the cost-effectiveness of this treatment. The aim of this study was to estimate the cost-effectiveness of TDF versus four oral nucleos(t)ide analogs [lamivudine (LAM), adefovir (ADV), telbivudine (LdT), and entecavir (ETV)] and from a pharmacoeconomic perspective to assess current drug pricing for TDF. METHODS: Based on Chinese healthcare perspectives, a Markov model was applied to simulate the lifetime (40-year time span) costs and quality-adjusted life-years (QALYs) for five different monotherapy strategies. Two kinds of rescue combination strategies (base-case: LAM + ADV then ETV + ADV; alternative: directly using ETV + ADV) were separately considered for treatment of patients refractory to monotherapy. Model parameters (including disease transition, cost, and utility) were obtained from previous Chinese population studies. Both branded and generic drugs were separately analyzed. Study model uncertainties were assessed by one-way and probabilistic sensitivity analyses. Two-way sensitivity analysis was used to explore uncertainties between efficacy and price of TDF. RESULTS: In the base-case analysis, the lowest lifetime cost and the best cost-effectiveness ratio were obtained by ETV, which was considered the reference treatment. LAM, ADV, and LdT treatments had significantly greater costs and lower efficacies. Compared to ETV, TDF was more effective but also more expensive. The incremental cost-effectiveness ratios of TDF versus ETV were much higher than the willing-to-pay threshold of $20,466 US dollars (USD) per QALY gained (3 × gross domestic product per capita of China, 2014). TDF would be the most cost-effective strategy if the annual cost did not exceed $2260 USD and $1600 USD for branded and generic drugs, respectively. CONCLUSIONS: For Chinese chronic hepatitis B patients, ETV is still the most cost-effective strategy over TDF and other nucleos(t)ide analogs, with a threshold of $20,466 USD/QALY gained.

Synthetic Toll-like receptor 7 ligand inhibits porcine reproductive and respiratory syndrome virus infection in primary porcine alveolar macrophages.

Porcine reproductive and respiratory syndrome virus (PRRSV), a common viral pathogen, causes huge annual economic losses to the swine industry worldwide. After triggering by specific ligands, the Toll-like receptor 7 (TLR7), a type of pattern-recognition receptor (PRR), induces antiviral cytokines production. Previously, we synthesized an adenine analog, designated SZU101, a TLR7-specific ligand. In this study, we assessed the inhibitory effect of SZU101 on PRRSV infection in vitro. SZU101 significantly suppressed PRRSV infection in primary porcine alveolar macrophages (PAMs) in a dose-dependent manner. Moreover, SZU101-induced inhibition involved NF-κB pathway activation in PAMs to initiate expression of TLR7-mediated cytokines and induce expression of downstream signaling IFN-stimulated genes (ISGs). Chloroquine, a TLR7 inhibitor, and BAY 11-7082, an NF-κB inhibitor, reversed both the SZU101-induced antiviral effect and induction of cytokine genes and ISGs expression. Therefore, SZU101 antiviral effects depend at least in part on TLR7-NF-κB signaling pathway. Additionally, administration of SZU101 enhanced the humoral and cell-mediated immune responses against PRRSV antigens in mice. Given these results, SZU101 holds promise as an antiviral agent and a vaccine adjuvant to prevent PRRSV infection in pigs.

Electroacupuncture improves memory and protects neurons by regulation of the autophagy pathway in a rat model of Alzheimer's disease.

BACKGROUND: Acupuncture is a potential therapy for Alzheimer's disease (AD), but its clinical effects and underlying mechanisms are not fully understood. Emerging evidence suggests autophagy is involved in ß-amyloid (Aß) clearance. We hypothesised that electroacupuncture (EA) treatment of AD involves the autophagy pathway in rats. METHODS: We injected 2µl Aß1-40 bilaterally into the hippocampi of 42 rats to establish AD. Rats remained untreated (AD group, n=14) or received 24 EA treatments at GV20+BL23 over 28 days from day 7 post-injection with/without co-treatment with 3-methyladenine (3-MA), an autophagy inhibitor (AD+EA+3-MA and AD+EA groups, respectively, n=14 each). Cognitive function was evaluated by Morris water maze (MWM) testing. Hippocampi were examined by transmission electron microscopy (TEM) and stained with haematoxylin and eosin/transferase dUTP nick end labelling (TUNEL) to assess neuronal morphology/apoptosis, respectively. Protein expression of Beclin-1, LC3 and Aß1-40 was examined. RESULTS: In the MWM test, the AD+EA group showed an improvement in parameters consistent with improved learning/memory compared to untreated AD rats, and 3-MA attenuated these effects. EA mitigated cellular apoptosis resulting from Aß infusion in the CA1 region and enhanced LC3II/LC3I ratios and Beclin-1 expression. Numerous autophagosome precursors and enlarged autophagosomes were observed by TEM in the hippocampi of EA-treated rats. Reduced Aß levels, and co-localisation of Aß and LC3II, were observed following EA treatment by immunofluorescence staining. EA+3-MA treated rats had much higher TUNEL-positive neurons, lower LC3II/LC3I ratios and Beclin-1 expression, and elevated Aß levels compared with EA alone. CONCLUSIONS: EA reduces neuronal apoptosis, enhances degradation of Aß, and improves learning/memory in AD rats by upregulating the autophagy pathway.

A quantitative multiplex nuclease protection assay reveals immunotoxicity gene expression profiles in the rabbit model for vaginal drug safety evaluation.

Any vaginal product that alters the mucosal environment and impairs the immune barrier increases the risk of sexually transmitted infections, especially HIV infection, which thrives on mucosal damage and inflammation. The FDA-recommended rabbit vaginal irritation (RVI) model serves as a first line selection tool for vaginal products; however, for decades it has been limited to histopathology scoring, insufficient to select safe anti-HIV microbicides. In this study we incorporate to the RVI model a novel quantitative nuclease protection assay (qNPA) to quantify mRNA levels of 25 genes representing leukocyte differentiation markers, toll-like receptors (TLR), cytokines, chemokines, epithelial repair, microbicidal and vascular markers, by designing two multiplex arrays. Tissue sections were obtained from 36 rabbits (6 per treatment arm) after 14 daily applications of a placebo gel, saline, 4% nonoxynol-9 (N-9), and three combinations of the anti-HIV microbicides tenofovir (TFV) and UC781 in escalating concentrations (highest: 10% TFV+2.5%UC781). Results showed that increased expression levels of toll-like receptor (TLR)-4, interleukin (IL)-1ß, CXCL8, epithelial membrane protein (EMP)-1 (P<0.05), and decreased levels of TLR2 (P<0.05), TLR3 and bactericidal permeability increasing protein (BPI) (P<0.001) were associated with cervicovaginal mucosal alteration (histopathology). Seven markers showed a significant linear trend predicting epithelial damage (up with CD4, IL-1ß, CXCL8, CCL2, CCL21, EMP1 and down with BPI). Despite the low tissue damage RVI scores, the high-dose microbicide combination gel caused activation of HIV host cells (SLC and CD4) while N-9 caused proinflammatory gene upregulation (IL-8 and TLR4) suggesting a potential for increasing risk of HIV via different mechanisms depending on the chemical nature of the test product.