The data presented set the stage for investigating both host-specific and virus-specific mechanisms that control primary and sequential DENV infections. Previous immunity is a major risk factor for dengue haemorrhagic fever, so these mice could potentially be used to study the role of cross-reactive sub-neutralizing antibodies and T cells during sequential DENV infections as well as to test drugs and

vaccines against dengue. Increased understanding of the contribution of host components to severe dengue disease selleck chemicals llc will lead to the development of effective therapeutics and vaccines. We thank Dr Alan L. Rothman for carefully reading this manuscript and Kim West for technical assistance. This project was supported by grant U19 AI57319 and U19 AI057234 from the National Institute of Allergy and Infectious Diseases, a grant from the Juvenile Diabetes Research Foundation and the Helmsley Foundation,

National Institutes of Health (NIH) grant CA34196, an NIH Diabetes Endocrinology Research Center (DERC) grant DK52530 and support from USAMRID. The authors declare no financial or commercial conflict of interest. “
“Control and termination of infection with Influenza A virus is associated with increased IL-10 production in mouse models. Notably, IL-10 can be produced by Treg. Therefore, we investigated whether the population of IL-10-producing influenza-specific CD4+ selleck inhibitor T cells comprised Treg as they are potent suppressors of the adaptive immune response. Influenza-specific IL-10-producing CYTH4 T cells were detected

in all human donors displaying influenza-specific immunity. Isolation of Matrix 1 protein-specific IL-10-producing T-cell clones revealed that a substantial proportion of these T-cell clones displayed the capacity to suppress effector cells, functionally identifying them as Treg. Both FOXP3+ and FOXP3− CD4+ Treg were isolated and all were able to exert their suppressive capacity when stimulated with cognate antigen, including influenza virus-infected cells. In vitro suppression was not mediated by IL-10 but involved interference with the IL-2 axis. The isolated Treg suppressed amongst others the IL-2 production of influenza-specific T-helper cells as well as partially prevented the upregulation of the high-affinity IL-2 receptor on CD8 effector cells. So far the induction of virus-specific Treg has only been studied in the context of chronic viral infections. This study demonstrates that virus-specific Treg can also be induced by viruses that are rapidly cleared in humans. CD4+ Treg can be generated both in the thymus and in the periphery 1. Generation of Treg in the periphery has been well demonstrated in mouse models 2–4. So far, pathogen-specific Treg have been isolated only in the context of chronic infections and viral-induced cancer in humans 5–8 and are thought to be the result of T-cell priming during chronic phases of disease.

Table 1 lists the primers that

were used for mRNA quantification. Samples were analysed using a Bio-Rad iCycler iQ (Bio-Rad, Hercules, CA). Changes in gene expression were determined by calculating the Δ cycle threshold (Ct) by subtracting the Ct for ribosomal protein L19 (RPL19) (reference gene) from the Ct of the gene of interest for each sample.26 The ΔCt of the control was subtracted from the corresponding treated sample giving rise to the ΔΔCt. The fold change was derived from the equation 2−[ΔΔ]Ct. To confirm that the reference gene ribosomal protein L19 was stably expressed in MoDCs and BDCs, a comparison was performed using either glyceraldehyde 3-phosphate dehydrogenase (GAPDH) or RPL19 as the Silmitasertib research buy reference gene. Similar trends in fold change were observed. Complementary DNA was diluted to generate

a standard curve whose correlation coefficient was > 0·99. The efficiency of qPCR was determined from the slope using the equation (10[−1/M] − 1) × 100 and ranged between 90% and 110%. To evaluate changes in cytokine secretion, 1 × 106 MoDCs or BDCs were incubated in 1 ml culture medium for 24-hr in six-well plates (Corning) and culture supernatants were collected. Concentrations of IL-6, MLN8237 clinical trial IL-8 and IL-10 were assayed using commercial kits as per the manufacturer’s instructions (R&D Systems, Minneapolis, MN). The ELISA for IFN-α, TNF-α and IL-12 were performed as previously described.27 Statistical analysis was performed by non-parametric Mann–Whitney U-tests (P-value < 0·05) using the statistical software programme graphpad prism 5 (GraphPad Software, Inc., La Jolla, CA). In this study, 800 ml of EDTA blood yielded approximately 2 × 109 PBMCs. Following CD14+ selection, an average of 2 × 108 monocytes were cultured in the presence of IL-4 and GM-CSF to Calpain generate MoDCs. On day 6, approximately 2 × 107 MoDCs were harvested and cultured for use. The CD14− population

was positively selected for cells expressing CD172, which equates to the BDC (CD14− CD172+) population. Approximately 3 × 107 BDCs were therefore isolated and rested overnight. In contrast to other studies, the protocol used in this study resulted in lower numbers of MoDCs compared with BDCs from an equal amount of blood.28 Dendritic cell morphology is characterized by a large cytoplasmic cell mass and extrusion of dendrites which increase the surface area available to sample and take up antigens. In this study, the morphologies of Giemsa-stained MoDCs (Fig. 1a) and BDCs (Fig. 1b) were compared. Both DC populations displayed a typical DC morphology, characterized by an irregular cell border with a large cytoplasmic cell mass. Expression of cell surface markers CD172, MHC II, CD16, CD1, CD80/86 and CD14 was assessed by flow cytometry in 6-day-old MoDCs and BDCs (Table 2). Both MoDCs and BDCs expressed all of these markers; however, BDCs showed similar expression of CD172 and MHC II, higher expression of CD16 and lower expression of CD80/86 and CD1.

In contrast, using Western blotting MAPK inhibitor in this study we found that TLR-4 expression specifically in AS T cells was suppressed by let-7i. As TLR-4 is expressed abundantly on monocytes, we proposed that the decreased expression of TLR-4 in AS T cells could be masked easily by the abundant amount of TLR-4 on monocyte or other cell types from AS patients. Moreover, in the cell transfection studies, we found that there were discrepancies between mRNA and protein expressions of TLR-4

due to the effect of let-7i (Figs 6 and 7). As TLR-4 is the prime cellular pattern recognition sensor for microbial pathogens, TLR-4 activation via LPS leads to production of proinflammatory Forskolin cytokines in innate immune systems [38]. Interestingly, TLR-4 is also expressed on T cells [39], which might have a different immunoregulatory

function in the adaptive immune system, as shown in our study. José et al. [33] have reported that LPS signalling through TLR-4 could suppress T cell receptor-dependent extracellular signal-regulated kinase 1/2 (ERK1/2) activation in CD4+ T cells in the murine model. Similar to their findings, in this study we demonstrated that LPS could exert an inhibitory signal on the T cell response in humans. Clinical observations revealed that there was a link between AS development with chronic prostatitis in men or pelvic inflammatory disease in women. It is purposed that the microbe infection is from a source of damage-associated molecular pattern molecules (DAMPs)

involved in AS pathogenesis. These DAMPs could activate TLRs to elicit the inflammatory reaction and ectopic enchondral bone formation in AS spine [32]. Although bacterial infection such as Chlamydia could cause chronic arthritis [40], it is still premature to conclude that bacterial infection can cause AS [41]. Conversely, evidence suggests that AS disease activity became worse, following the different bacterial infections such as Salmonella, Yersinia, Campylobacter and Chlamydia [42-46]. Although molecular mimicry between the bacterial components and self-peptides was considered to play a role [47], our results may provide an alternative explanation, that the bacterial LPS could suppress Ergoloid IFN-γ production in activated normal T cells. However, this regulatory mechanism was abrogated by the over-expressed let-7i in AS T cells (Fig. 8a). IFN-γ is a key proinflammatory cytokine which has been shown to be elevated in serum from AS patients [48]. Although we found no correlation between let-7i and the mRNA expression of IFN-γ in AS patients (Fig. 9b), contradictory to the finding that let-7i may regulate IFN-γ production (Fig. 8b) it is possible that various factors, such as viral or bacterial infection, trigger IFN-γ gene expression to confound our results.

The PCR samples (100 μl final volume) contained 5 μl cDNA, 0·2 mm dNTPs, 1·5 mm MgCl2, 20 pmol Igκ-5′ primer, 10 pmol Igκ-3′ primer, and 0·5 μl Taq polymerase (5 U/μl) (Invitrogen). Primer sequences are provided in Supplementary material, see Table S1. Thermal cycling conditions were as follows: 94° for 1 min; 60° for 2 min and 72° for 2 min for 30 cycles, followed by a final extension at 72° for 6 min. Amplified cDNAs were cloned using the TOPO-TA cloning kit (Invitrogen),

and individual clones were sequenced. To identify Vκ segment usage in the cloned cDNA, the NCBI database was queried using IgBLAST. Cohorts of 8-week-old wild-type and dnRAG1 mice were immunized with the hapten NP (4-hydroxy-3-nitrophenylacetyl) conjugated to either chicken gamma-globulin (NP-CGG; Biosearch Technologies, Novato, CA) or aminoethylcarboxylmethyl-FICOLL (NP-AECM-FICOLL; click here Biosearch Technologies), essentially as described elsewhere.25 To prepare the immunogen, NP-CGG or NP-Ficoll (100 μg) was dissolved

in 10% aluminium potassium sulphate and precipitated by adjusting the pH to 6·2 with 1 m potassium hydroxide. Alum precipitates were washed three times with PBS, and resuspended selleckchem in 200 μl PBS. Wild-type and dnRAG1 mice were injected intraperitoneally with either NP-CGG or NP-Ficoll. Some animals received a booster injection of antigen at day 7 (10 μg intravenously). Animals receiving no injection to or alum only served as controls. Levels of NP-specific antibodies were measured by ELISA in peripheral blood collected at day 7 (primary) or day 21 (secondary). Serum IgM and IgG levels were quantified using a commercially available sandwich ELISA according to the manufacturer’s instructions (IMMUNO-TEK mouse IgM and IgG immunoglobulin ELISA kit; ZeptoMetrix, Buffalo, NY). The NP-specific antibodies were detected as described by von Bulow et al.26 Optical density was measured at 450 nm using the GENios ELISA plate reader running the Magellan reader

control and data reduction software (Tecan Austria Gmbh). To generate dnRAG1 mice, we prepared a construct containing a RAG1 cDNA encoding a full-length catalytically inactive form of RAG1 under the transcriptional control of an H-2kb promoter, a genomic fragment of the human β globin gene to provide RNA splice donor sites and a polyadenylation signal, and an immunoglobulin heavy chain enhancer element (IgH Eμ) (Fig. 1a). RAG1 expressed from this construct lacked an epitope tag to avoid potential tag-associated artefacts that could alter RAG protein localization, regulation, or activity. Previous studies have shown that this promoter–enhancer combination supports transgene expression in the B-cell and/or T-cell lineage in founder-specific manner.9 Using PCR and Southern blotting approaches to screen founder lines (Fig.

Due to differences in dietary fats in the western world in the United States versus Europe [22], it is likely that the diet-induced changes in intestinal microbiota composition could partly explain the controversy regarding, e.g. the Firmicutes/Bacteroidetes ratio in humans [4, 14]. Nevertheless, PF-02341066 molecular weight it is now accepted that intestinal microbiota are involved in obesity, as germ-free ob/ob mice on both normal chow and high-fat diets remain

significantly leaner than conventionally raised mice, despite a significantly higher food intake [23]. In line with this, metagenomic sequencing of the caecum microbiome of these ob/ob mice revealed that an enrichment of genes was involved in the breakdown

of complex dietary polysaccharides [18]. Similar alterations showing enriched bacterial genes involved in carbohydrate sensing and degradation have also been observed in obese humans [24]. Studying intestinal microbial composition in well-phenotyped human subjects enrolled in relatively large metagenome-wide association studies (MGWAS) in both Chinese and European populations has further increased our understanding of the gut microbiota in the development of obesity and insulin resistance [25-27]. Karlsson et al. detected an enrichment of L. gasseri and S. mutans (both RO4929097 mouse commensal bacteria in the mouth and upper intestinal tract) to predict development of insulin resistance in their cohort of postmenopausal obese Caucasian females [26]. Conversely, Qin et al.’s Chinese T2DM cohort demonstrated that Escherichia coli, a Gram-negative check details bacterium which is associated with development of low-grade endotoxaemia, was more abundant. Moreover, clusters of genomic sequences acted as the database signatures for specific groups of bacteria and both studies found independently that subjects with T2DM were characterized by decreased

short chain fatty acid (SCFA) butyrate-producing Clostridiales bacteria (Roseburia and F. prausnitzii), and greater amounts of non-butyrate producing Clostridiales and pathogens such as C. clostridioforme, underscoring a potential unifying pathophysiological mechanism. It has long been recognized that insulin resistance and development of type 2 diabetes are characterized by systemic and adipose inflammation [19, 28]. The lipopolysaccharides (LPS) produced in the intestine due to the lysis of Gram-negative bacteria triggers proinflammatory cytokines that result in insulin resistance both in mice [5] and humans [29]. A more causal role was defined when germ-free mice were colonized with E. coli, as this promoted macrophage accumulation and up-regulation of proinflammatory cytokines resulting in low-grade inflammation [30].

There was a relation between fungal exposure at home and the spontaneous PBMC secretion of IL-6, IL-10 and IL-12 among subjects with sarcoidosis. A significant relationship was observed between disease severity, measured as chest X-ray scores indicating granuloma infiltration, and the P-glucan- and LPS-induced secretion of all cytokines. There was also a positive relation between the P-glucan-induced secretion of IL-12 and the duration of symptoms. There are some limitations to the study. The FCWA check details and LPS preparations used in the study were chemically well-defined compounds of bacterial and fungal origins but these are not wholly representative of the agents as present in the environment [15]. S-glucan and P-glucan

were purified from Alcaligenes faecalis, but in nature β-glucan is present together with capsular materials and chitin. The chitin preparation used was a de-acetylated form of chitin. LPS is a chemically purified lipopolysaccharide from Gram-negative bacteria, whereas the endotoxin present in nature also comprises proteins and sugars from the cell wall of Gram-negative bacteria [22].

In view of these differences between the substances used in the PBMC stimulation experiments and natural agents, caution should be applied in the interpretation of the in vitro findings and their relevance for clinical conditions. If, on the other hand, observations from exposures and cytokines in vivo parallel the in vitro results, the validity of the latter is supported. The in vitro method used also has some limits in terms of interpretation. A potential shortcoming https://www.selleckchem.com/products/gsk2126458.html is the lack of definition of different cell types. Due to the chronic inflammation subjects with sarcoidosis might have a different cell population particularly regarding lymphocytes, both in numbers and subtypes. Thus differences in cytokine production between patients with sarcoidosis and controls could be due to different proportions of responsive cells in the PBMC isolates. PBMC consist, however, mainly of monocytes and lymphocytes and the proportion reflects the monocyte/lymphocyte proportion

in white blood cells. These were counted in all our sarcoidosis stiripentol patients and only minor changes were present in the mono/lymph ratio compared to controls. From a clinical viewpoint, the presence of an inflammation is the most important issue for the patient. Whether or not this is due to a different distribution of cells is interesting from a mechanistic point of view, but not for the patient. The conclusion that subjects with sarcoidosis react more to FCWA and to the fungal exposure at home is thus a relevant finding, irrespective of the underlying mechanism. The results confirm findings from many previous investigations where FCWA were found to have important immunomodulating characteristics [14]. The FCWA used here had different effects on the secretion of cytokines from PBMC. P-glucan induced a high secretion of all cytokines.

Overall, studies with internal controls were limited and loss to follow up was high. The average decrement in GFR (22 studies) in donors with normal renal function after donation was 26 mL/min per 1.73 m2 (range 8–50). After 10 years (8 studies), 40% (range 23–52%) of donors had a GFR between 60 and 80 mL/min per 1.73 m2, 12% (range 0–28%) had a GFR between 30 and 59 mL/min per 1.73 m2 and 0.2% (range 0–2.2%) had a GFR less than 30 mL/min per 1.73 m2. In the 6 controlled studies where average follow up was at least 5 years, the

post-donation weighted mean difference in GFR among the donors compared with controls was −10 mL/min per 1.73 m2 BMN 673 nmr (95% CI: 6–15). Garg and colleagues note no evidence of an accelerated loss of GFR over that anticipated with normal ageing with the lower absolute GFR being attributable to the decrement occurring Erismodegib mw as a result of nephrectomy. However, they also note that the prognostic significance of the reduced GFR in healthy donors is unknown given the mechanism of reduction is different to that which occurs in CKD. The evidence with respect to the outcome of living kidney donors who have reduced GFR at the time of donation is limited. A systematic review and meta analysis of health outcomes for living donors with isolated medical abnormalities including age, obesity, hypertension or antihypertensive medication, haematuria, proteinuria, nephrolithiasis and reduced GFR (defined as ≤80 mL/min) has been recently completed by

Young et al.1 Only one study was identified that compared donors with a reduced GFR (n = 16) with those having normal GFR (n = 75).21 This was also the Monoiodotyrosine only study identified that considered proteinuria as an IMA. Although this was a prospective study, the proportion lost to follow up was not reported. One year after donation, the GFR was lower in the IMA group (51.7 ± 11 mL/min) compared with the control (68.0 ±  15 mL/min).

At follow up 8 years after nephrectomy, the donor with the lowest GFR at 1 year (44 mL/min) had a GFR of 63 mL/min. Young and colleagues also note that there are very few studies documenting important health outcomes among living kidney donors with IMAs. Across all IMA groups, longer term assessments (≥1 year) of blood pressure, proteinuria and renal function have been reported in only 3, 2 and 10 studies, respectively. Only 17 of the 37 studies included prospective data. A limited number provided loss to follow up and the studies were small. Overall, the ability of the primary studies to identify significant differences in long-term medical risks, including long-term renal function is limited.1 In the study by Rook et al. examining the predictive capacity of pre-donation GFR, 31 of 125 donors had a post-donation GFR < 60 mL/min per 1.73 m2.7 In this group, the mean pre-donation GFR measured by iothalamate was 99 mL/min ± 12 mL/min (88 ± 10 mL/min per 1.73 m2), while the pre-donation CG GFR was 83 ± 21 mL/min and the pre-donation GFR by simplified MDRD was 69 ± 8 mL/min.

Data are expressed as mean±SD for each group. Statistical differences between groups were evaluated using a Mann–Whitney test using GraphPad Prism 4.03 software. p<0.05 was considered statistically significant. We thank Dr. Randle Ware for critical reading of the see more manuscript and the members of the Laboratory of Autoimmunity for their help. This work was supported

by the National Institutes of Health grant RO1AI052227, MSNRI and DNRG to V.K. Conflict of interest: The authors declare no financial or commercial conflict of interest. “
“One of the major obstacles in dissecting the mechanism of pathology in human primary biliary cirrhosis (PBC) has been the absence of animal models. Our laboratory has focused on a model in which mice, following immunization with a xenobiotic chemical mimic of the immunodominant autoepitope of the E2 check details component of pyruvate dehydrogenase complex (PDC-E2), develop autoimmune cholangitis. In particular, following immunization with 2-octynoic acid (a synthetic chemical mimic of lipoic acid-lysine located within the inner domain of PDC-E2) coupled to bovine serum albumin (BSA), several strains of mice develop typical anti-mitochondrial autoantibodies and portal inflammation. The role of innate immune effector cells, such as natural killer (NK) cells and that NK T cells, was studied in this model based on the hypothesis that early events during

immunization play an important role in the breakdown of tolerance. We report herein that, following in-vivo depletion of NK and NK T cells, there is a marked suppression of anti-mitochondrial autoantibodies and cytokine production from autoreactive T cells. However, there

was no change in the clinical pathology of portal inflammation compared to controls. These data support the hypothesis that there are probably multiple steps in the natural history of PBC, including Aurora Kinase a role of NK and NK T cells in initiating the breakdown of tolerance. However, the data suggest that adaptive autoimmune effector mechanisms are required for the progression of clinical disease. Primary biliary cirrhosis (PBC) is an autoimmune disease of the liver characterized by specific destruction of the small bile ducts and the presence of readily detectable levels of anti-mitochondrial antibodies (AMA) [1–3]. Recently, we reported that natural killer (NK) cells are involved in the destruction of cholangiocytes and NK T cells are partly responsible for the exacerbation of disease in PBC [4–6]. While these data are consistent with the view that innate immune effector mechanisms serve as a bridge to acquired immunity, and the data imply a major role for innate immune effector mechanisms in the initiation of pathogenesis of human PBC [7,8], the precise details of how such innate immune effector mechanisms influence the generation of pathogenic acquired immune responses remains poorly understood.

This study compared the adhesive and chemotactic functions of neutrophils from RA patients in activity (DAS28 > 3.2) and not in activity (DAS28 < 2.6) and observed the effects of different treatment approaches on these functions. Neutrophils were isolated from healthy controls (CON), and patients with active or inactive RA in use of therapy not specific LBH589 research buy for RA (NSAIDs), in use

of DMARDs and in use of anti-TNF-α therapy. Adhesive and chemotactic properties were evaluated using in vitro assays; adhesion molecule expression was assessed by flow cytometry and real-time PCR and circulating chemokines were determined by ELISA. No significant alterations in the adhesive and chemotactic properties of neutrophils from active RA were observed when compared to CON neutrophils, independently of treatment regimen. In contrast, neutrophils from RA patients in disease remission presented

reduced adhesive properties and a lower spontaneous chemotactic capacity, in association with decreased adhesion molecule expression, although profiles of alterations differed for those patients on DMARDs and those on anti-TNF-α therapy. Circulating levels of the major neutrophilic chemokines, IL-8 and epithelial neutrophil activating peptide-78, were also significantly INCB024360 nmr decreased in those patients demonstrating a clinical response. Remission of RA appears to be associated with ameliorations in aspects important for neutrophil adhesion and chemotaxis; whether these alterations contribute to decrease neutrophil migration to the synovial fluid, with next consequent improvements in the clinical manifestations of

RA, remains to be determined. Rheumatoid arthritis (RA) is a common systemic autoimmune disease, characterized mainly by synovial hyperplasia and symmetric polyarticular joint disorders [1]. Although the pathophysiology of this disease is not fully understood, it is known that the chronic inflammatory nature of the disease causes the migration of leucocytes from the peripheral circulation into the synovial tissue and synovial fluid (SF) via their interaction with endothelial cells, cellular adhesion molecules, cytokines, chemokines and receptors. The synovial tissue of RA individuals becomes replete with mononuclear cells [2, 3] while the neutrophils constitute over 90% of cells in the SF [4]; these phenomena lead to the proliferation of fibroblast-like synoviocytes with consequent destruction of cartilage and bone [5]. The migration of neutrophils to inflammatory foci is thought to be initiated by the capture, rolling and subsequent firm adhesion of the cells on the endothelium in response to chemotactic molecules such as the CXC chemokines interleukin (IL)-8 and epithelial neutrophil activating peptide (ENA)-78 [6].

[147-151] We would like to believe that in the near future TAM-targeted strategy will be clinically accepted as a valuable adjuvant therapy for SB203580 concentration cancer patients. However, we have come to appreciate the fact that cancer is a systemic disease and TAMs are involved in tumour progression through rather complex mechanisms. TAM-targeted therapy, therefore, requires an overall understanding about TAM functions in tumour development. One major gap in our knowledge is why TAM infiltration is associated with poor prognosis in many types of

cancers but with favourable survival in others. Although a few pieces of evidence indicate the micro-anatomical location and macrophage phenotype might be responsible for this dichotomy,[152-154] clinical evidence is substantially lacking. Second, it would be interesting to identify TAM-specific molecules

that could serve buy LDE225 as targets for tumour therapy, because previous identified factors (e.g. VEGF, MMPs, TGF-β and CXCL-12) important for TAM-mediated tumour progression,[3, 4, 7-9, 75] are also produced by cancer cells themselves. Hopefully, recent clinical and experimental investigations have identified several tumour-promoting molecules (e.g. CCL-18 and IRAK-M) predominantly produced by M2 TAMs.[155, 156] Third, what should not be neglected is the close interaction between macrophages and other stromal cells within the tumour microenvironment. A better understanding of those connections will contribute to TAM-targeted adjuvant Bay 11-7085 therapies. The fourth inherent issue is how to keep the balance between ‘cancer-inhibiting inflammatory responses’ and ‘cancer-promoting inflammatory

responses’.[157, 158] More biological understandings and pharmacological approaches are needed to fill this gap of our knowledge. Furthermore, a practical issue for developing TAM-targeted therapy is that, clinically, how should a drug be administered at the right time and to the right place so that the tumour-promoting TAMs could be depleted or re-educated whereas the tumoricidal macrophages in tumours or healthy tissues remain unaffected. In summary, more comprehensive understanding of the properties of TAMs and their interactions with the tumour microenvironment, together with advances in diagnostic/therapeutic techniques, will be required to facilitate the development and clinical application of TAM-targeted adjuvant cancer therapies. Our deepest gratitude goes first and foremost to Dr Meiyi Pu for her critical reading of the manuscript and her great contribution to the English improvement. Without her help, this article could not have reached its present form. We also thank Dr Changhua Zou for her wonderful suggestion. This work was supported by a grant from the West China Hospital of Sichuan University (Huaxi Grant 13708002). The authors declare having no conflicts of interest. “
“Viral diversity is a challenge to the development of a hepatitis C virus (HCV) vaccine.