The individual PD20FEV1 × 10 was then used for the
subsequent Segmental Allergen Provocation (SAP). Inhaled and segmental allergen challenges were separated by at least 4 weeks. Bronchoscopy was performed as previously described [29, 42]. A volume of 2.5 ml of 0.9% saline was instilled into the anterior basal segment of the left lower lobe (B8 left) and one of the segments of the lingula (B4 or B5 left). Allergen diluted in 2.5 ml of saline was instilled into the anterior basal segment of the right lower lobe (B8 right) and the medial or lateral segment of the right middle lobe (B4 or B5 right). After 10 min, Erastin mouse bronchoalveolar lavage was performed in the anterior basal segments of the right and left lower lobes. Patients were re-bronchoscoped Panobinostat cost at different time points: In the first group, the second lavage was performed after 18 h in segments B4 or B5 right and left. Some of these patients also participated in the second part of the trial. This second group was lavaged 10 min and 42 h after segmental allergen challenge (Table 1). In the third arm of the trial, four patients were
lavaged 10 min and 162 h after allergen challenge. In patients who participated repeatedly the segmental allergen challenges were separated by at least six months. In all patients, peripheral blood was taken before bronchoscopy. From seven healthy subjects and seven patients with allergic asthma, 250-ml whole blood was drawn and mixed well with heparin. Cell subtypes were separated by Ficoll centrifugation. PBMC-CD14+ were harvested, and after washing and counting monocytes were separated via immunomagnetic BCKDHA separation by AutoMACS system (Miltenyi Biotec GmbH, Germany) after labelling with CD14 antibody (Miltenyi Biotec GmbH). CD14+
monocytes were washed; purification was controlled by flow-cytometry (94–98% purified monocytes, contamination with lymphocytes was <2%), and 5 × 105 cells per well were cultured in 1 ml RPMI 1640 medium (GIBCO, Paisley, Scotland, UK) + 10% FCS (Seromed, Berlin, Germany) + 1% penicillin/streptomycin (Biochrom AG, Berlin, Germany) at 37 °C and 5% CO2. Cells differentiated to macrophages in about 5 days. Medium was exchanged every 2 days. The above-mentioned PBMC-CD14+ cells (5 × 105 cells in 1 ml) were stimulated with either human IL-17 (50 ng/ml), LPS (10 ng/ml), leukotriene D4 (LTD4) (10−11 M) or a combination of LPS and LTD4 for a duration of 6, 12 and 24 h. Cells were also stimulated with LTD4 in the presence of the leukotriene antagonist Montelukast. LTD4 was added to cell cultures 30 min after stimulation with Montelukast (10−11 M), and cultures were incubated for 6, 12 and 24 h. Cell culture supernatants were stored at −20 °C until sCD14 measurement with an ELISA kit (IBL Hamburg, Germany) according to the manufacturer’s instructions. Data were analysed by SPSS software package. Results are reported as median (range) or as single values and median (Figs. 2–5).
[14, 15] Heart failure may develop ‘de novo’ after receiving a kidney transplant. Using United States Medicare Claims data, the cumulative incidence of de novo CHF was 10.2% after 12 months and 18.3% after 36 months compared with 12.0% and 32.3%, respectively for patients remaining on dialysis on the transplant waiting list. The cumulative incidence of de novo CHF in patients who survived the first post-transplant year without CHF
has been reported to be 3.6% at 5 years and 12.1% at 10 years. The objectives of this guideline are to summarize the available evidence for the treatment of CHF in patients with CKD defined by a GFR < 60 mL/min not requiring dialysis, patients receiving dialysis and kidney transplant recipients. The following treatments have been FK506 considered: Blockade of the renin-angiotensin system Blockade of beta-adrenergic receptors Aldosterone antagonists BYL719 research buy Digoxin Vasodilators
(hydralazine and nitrates) Treatment of anaemia Strategies to control volume state Use of Implantable Devices Other therapies The recommendations for patients with CKD and kidney transplant are grouped together because these patients are similar in terms of current actual kidney function, and there are no trials that specifically enrolled kidney transplant recipients with CHF to study a heart failure intervention. It is acknowledged that kidney transplant recipients will differ in many ways from CKD, including time receiving dialysis, presence of arteriovenous fistula and immunosuppression. A number of RCTs have been performed in patients with CHF that provide a strong evidence base underpinning many guideline recommendations for the general population.[6, 18, 19] The recommendations for patients with CKD are based
on post-hoc analyses of RCTs of therapies in patients with heart failure. Although these are post-hoc analyses, a large proportion of patients in these studies had an eGFR < 60 mL/min per 1.73 m2 so the results of these trials can be applied to CKD Stage 3. However, PDK4 fewer patients in the trials had an eGFR < 30 mL/min per 1.73 m2 so this should be borne in mind when applying these guidelines to such patients. There are no data specifically for kidney transplant recipients but it is considered reasonable to apply the CKD recommendations to this group, acknowledging this lack of specific data. For dialysis patients, there are smaller trials of lower quality but these data are generally consistent with the CKD and general population data. *Explanation of grades The evidence and recommendations in this KHA-CARI guideline have been evaluated and graded following the approach detailed by the GRADE working group (http://www.gradeworkinggroup.org). A description of the grades and levels assigned to recommendations is provided in Tables 1 and 2. High quality of evidence. We are confident that the true effect lies close to that of the estimate of the effect. Moderate quality of evidence.
The second strategy, developed mainly over the past decade, consisted of more ambitious forms of immune therapy
not aiming at immunosuppression but at inducing/restoring self-tolerance buy MK-2206 to well-defined β cell antigens. The rationale was based on the well-established notion that antigen delivery depends upon the molecular form of the antigen and its route of inoculation, and may lead either to effective immunization or to immune tolerance. This concept stemmed from pioneering experiments performed by D. W. Dresser in the early 1960s, showing that heterologous immunoglobulins that are immunogenic if administered in aggregated form induce specific unresponsiveness/immune tolerance, or ‘immune paralysis’, if injected intravenously (i.v.) in non-aggregated form . Thus it made sense to use well-defined autoantigens as therapeutic tools to attempt inducing/restoring self-tolerance in T1D. As in many other autoimmune diseases, in T1D various candidate autoantigens have been incriminated as potential triggers and targets of the disease. These include the main β cell hormone proinsulin/insulin itself, glutamic acid decarboxylase (GAD), a β cell-specific protein
phosphatase IA-2, a peptide (p277) of heat shock protein 60 (hsp60), the islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP), a preferential
target of pathogenic CD8+ T cells, and the most recently characterized zinc transporter this website ZnT8. Targeting some of these antigens has proved successful in NOD mice, as disease was effectively prevented by administration of protein or specific peptide antigens such as pro-insulin, insulin, GAD, the p277 peptide of hsp60 using various routes [i.v., subcutaneous (s.c.), oral, intrathymic, intranasal]. Although highly effective in the experimental setting, the transfer to the clinic of β cell autoantigen-induced strategies was beset by a number of difficulties. Antigens used in patients included insulin or altered insulin peptides, GAD65 and the hsp60 Bumetanide p277 peptide (DiaPep277). Most applications have been via administration of the antigen or peptide alone, and one approach has included the administration of antigen plus adjuvant. Insulin has been the main antigen used clinically. It was readily available for clinical use; experiments in animal models consistently showed effects in preventing diabetes; and several evidences suggested that insulin could be a primary autoantigen in T1D. Insulin has been used as an immunotherapy via s.c., i.v., oral and intranasal routes. Two trials performed after diabetes onset in approximately 100 patients have tested the use of oral insulin at a limited dose range without observing efficacy [21,22].
[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 selleck chemical 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 click here 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 Aspartate 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.
Since previous studies have shown that iNKT17 cells can secrete IL-17 through TCR engagement 20, we investigated whether CD1d was HSP assay required for IL-17A mRNA
expression by iNKT17 cells in the pancreas (Fig. 3E). To address this question, we used Vα14 NOD mice expressing CD1d solely in the thymus (CD1dpLck Vα14 NOD mice) 31. RORγt, IL-23R and IFN-γmRNA expression was similar in pancreatic iNKT cells from both types of mice. However, IL-17A mRNA expression was significantly decreased (3-fold) in iNKT cells from mice lacking peripheral CD1d expression. Altogether, our data suggest that iNKT17 cells are activated locally in the pancreas in a CD1d-dependent manner. To evaluate the role of iNKT17 cells in type 1 diabetes, we reconstituted immunodeficient NOD mice with different iNKT cell subsets and analyzed the induction of diabetes after transfer of anti-islet BDC2.5 T cells 32. Since there is no specific antibody available to purify iNKT17 cells, we first determined the frequency of iNKT17 cells in different iNKT cell subpopulations divided according to CD4 and NK1.1
expression of donor cells. As shown in Fig. 3A and Supporting Information Fig. 2, iNKT17 cells are mainly present in the CD4− iNKT cell population and at a higher frequency among NK1.1− CD4− iNKT cells. Therefore, we enriched iNKT17 cells based on their lack of CD4 expression and they were found to represent around 23% of the injected CD4− iNKT cell population (Fig. 3B). Recipient NOD mice were reconstituted buy SAR245409 with CD4− or CD4+ iNKT cells, which were detected in pancreas before BDC2.5 T-cell transfer (Fig. 3B). In order to detect an eventual pathogenic role of iNKT17 cells, all recipient mice were injected with a low number of BDC2.5 T cells, which induces around 30% of diabetes in control mice devoid of iNKT cells (Fig. 3C). Interestingly, in the group of mice reconstituted with CD4− iNKT cells, the incidence of diabetes was significantly (p=0.036) increased Quinapyramine and reached 70%. In contrast, reconstitution with CD4+ iNKT
cells significantly (p=0.033) prevented the development of diabetes. Moreover, when CD4− iNKT cells were further divided according to NK1.1 expression, only NK1.1− CD4− iNKT cells containing the higher frequency of iNKT17 cells exacerbated diabetes (Fig. 3D). Since diabetes induced by diabetogenic BDC2.5 T cells is associated with their production of IFN-γ 13, we have analyzed whether the presence of iNKT cell subsets have influenced their production of IFN-γ and IL-17. As previously described 13, in diabetic control mice devoid of iNKT cells, BDC2.5 T cells produced large amount of IFN-γ in both PLNs and pancreas (Fig. 4A). In diabetic mice reconstituted with CD4− iNKT cells, production of IFN-γ by BDC2.5 T cells was similar as in diabetic control mice and production of IL-17 remained low, less than 1%. While cytokine production by BDC2.5 T cells was similar in both groups of mice, the frequency of BDC2.
, Shanghai, China) and stimulated with HspX, Ag85B, purified protein derivative CSF-1R inhibitor and Mpt64190–198, respectively, with ConA and PBS as positive and negative controls, for 36 h at 37 °C, 5% CO2. The cells were then removed, and 200 μl/well ice-cold deionized water was added to lyse the remaining cells. The plates were incubated on ice for 15 min, after which they were washed 10 times with PBST. Next, biotinylated detector antibody solution was added and the plates were incubated
for 1 h at 37 °C. The plates were washed five times with PBST, after which 100 μl/well streptavidin–horseradish peroxidase was added. The plates were again incubated for 1 h at 37 °C and washed five times with PBST. One hundred microlitres of AEC (3-amino-9-ethylcarbazole) substrate was added to each well. The plates were developed for 25 min at room temperature in the dark. The wells were washed with distilled water to stop development when the stained cells were counted on an automated ELISPOT reader and analysed with ImmunSpot software (Bio-sys, GmbH, Karben, Germany). Protective
efficacy assay. Mice were sacrificed for bacterial CFU count at 6th week post-challenge with H37Rv. The lower left lobe of the lungs from infected mice (n = 7) was harvested, homogenized in 0.05% PBS-Tween 80 and planted in 10-fold dilutions (10–1000) GSK3 inhibitor on Middlebrook 7H11-OADC agar (BD, Franklin Lakes, NJ, USA) containing ampicillin (10 μg/ml) to prevent contamination. Bacterial colonies were counted 3 weeks after incubation CYTH4 at 37 °C. Histopathology of the lung tissues. Each upper lobe of the left
lung of infected mice (n = 5) was harvested 6 weeks after challenge. The lobes were fixed with 10% neutral buffered formalin. After 2 weeks, each lobe was bisected with 5 μm thick to examine the same area of the lung in all mice. The sections were stained with haematoxylin and eosin (HE) and Ziehl–Neelsen Method. Granulomas area was divided by total section area to determine the affected area in a section. Histopathology was evaluated by three pathologists independently. Statistical analysis. The results were expressed as means ± standard deviation (SD) and analysed by SPSS10.0 software (Statistical Product and Service Solutions Company, Chicago, IL, USA). The significance of differences among the groups was determined by analysis of variance (anova). Independent-samples t-test was used for Ziehl–Neelsen stain. Probability values (P < 0.05) were considered as statistically significant. The correct DNA sequence for the recombinant fusion protein, AMH was confirmed by sequencing and was found to encode a protein with molecular weight of 54.6 kDa. AMH was overexpressed in E. coli in inclusion bodies, which were subsequently dissolved and purified with Ni-NTA His affinity chromatography.
In vitro experiments have revealed that DMF, as well as its primary metabolite monomethyl fumarate (MMF), can exert immunomodulatory effects on T-cell subsets as well as on antigen-presenting cells,[93, 94] and experiments in EAE have demonstrated that DMF is effective in
both preventive HM781-36B order and therapeutic applications, albeit marginal in chronic EAE, promoting myelin and axonal preservation and reducing astrocyte activation.[95, 96] It has been speculated that part of the effect of DMF could be mediated through modulation of microglia phenotype. Histological studies demonstrated that, during the acute phase of EAE, Mac-3-positive cells (microglia and macrophages) are significantly reduced in the spinal cord of DMF-treated animals. Such an observation is also supported by in vitro studies in which pre-treatment with DMF can inhibit LPS-induced activation of microglial cells by reducing
the expression of NO, TNF-α, IL-1β and IL-6, possibly through an inhibition of the extracellular-signal regulated kinase pathway and an activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. While in vitro data prompted the hypothesis that DMF and MMF could affect microglia activation through Nrf2, Selleckchem Cisplatin a pathway involved in the expression of proteins critical in the detoxification of reactive oxygen and reactive nitrogen species,[97, 98] this has not been demonstrated in vivo. Indeed, although Linker et al. showed
that Nrf2 is required for the therapeutic effect of DMF, double-labelling much of Nrf2 with a marker for microglia did not reveal an increase of its expression in those cells after DMF treatment in EAE-affected mice. Further in vitro and in vivo studies are needed to dissect the pathways through which DMF promote an alternative neuroprotective phenotype in microglia. Mesenchymal stem cells (MSC) are currently being investigated as an alternative therapeutic approach for MS. The potential therapeutic use of MSC for neurodegenerative diseases was originally considered as related to their possible regenerative function through their ability to differentiate into mesodermal tissues and perhaps into other embryonic lineages. However, recent observations have indicated that, upon systemic administration, most MSC are rapidly entrapped in the lungs, and only a few engraft into injured CNS, where they display negligible transdifferentiation capacity.[100-102] In vitro studies demonstrating that MSC can modulate several effector functions of cells of both the adaptive and innate immune systems introduced the possibility that MSC might be effective in EAE. Indeed, Zappia et al.
Disease is initiated by autoreactive T cells, which escape negative selection by expressing a second TCR with a different specificity or an altered affinity. Transfer of Ag-specific Treg cells ameliorates the early onset signs of disease but does not prevent the development of long-term chronic pathologies. Altogether, our results suggest that Ag dose directly affects Treg-cell generation and thus, the set-up of T-cell tolerance. “
“The NOD-like receptor (NLR) family pyrin domain-containing 3 (NLRP3) inflammasome is a cytoplasmic protein complex that mediates inflammatory
responses to a broad array of danger signals. The inflammasome drives caspase-1 activation and promotes secretion of the pro-inflammatory cytokines IL-1β and IL-18, and might also participate in other cellular processes. Here, we tried to identify new pathways regulated by the NLRP3 inflammasome in murine dendritic cells (DCs) Quizartinib mouse in response to monosodium urate (MSU) crystals. Using a transcriptomic approach, we found that DCs from Nlrp3−/− mice responded to MSU with differential expression of genes involved in the DNA damage response and apoptosis. Upon exposure to see more MSU or other ROS-mobilizing stimuli
(rotenone and γ-radiation), DNA fragmentation was markedly ameliorated in Nlrp3−/− and casp-1−/− DCs compared with WT DCs. Moreover, Nlrp3−/− DCs experienced significantly less oxidative DNA damage mediated by ROS. A significant decrease of the expression of several genes involved in double-strand and base-excision DNA repair was observed in WT DCs. Basal DNA repair capacity in WT DCs resulted in activation and stabilization of p53 in vitro and in vivo, which resulted 4��8C in increased cell death compared with that in Nlrp3−/− DCs. These
data provide the first evidence for the involvement of the NLRP3 inflammasome in DNA damage responses induced by cellular stress. Multicellular organisms are constantly exposed to environmental assaults and have evolved several mechanisms that either promote cellular repair or induce cell death in order to maintain tissue integrity. In particular, the immune system has evolved specialized innate cells that mediate recognition of invading microbes and host perturbations to initiate a potent set of defense mechanisms. To this end, innate cells are equipped with a range of surface and intracellular receptors that recognize both microbial-associated molecular patterns and danger-associated molecular patterns (DAMPs). When damage is not repairable, the damaged cells die and release a multitude of poorly defined DAMPs, which in turn elicit an inflammatory response. Inflammation can be both good and bad, depending on the situation. The NOD-like receptor (NLR) family pyrin domain-containing 3 (NLRP3) inflammasome is a multiprotein complex, which can drive inflammatory responses by promoting the release of IL-1β and IL-18 from innate cells .
Those authors hypothesized that a state of unresponsiveness to the endogenous microflora may be apparent only after a transient mucosal immune response has occurred . The response to bacteria and bacterial antigens we observed in our experiment might be elevated due in part to a transient unphysiological high load of bacteria in the axenic mice; however, it might mimic a response that occurs on a frequent basis, albeit less pronounced,
whenever a new bacterial strain is introduced in the intestinal lumen. The changes in the intestinal milieu with regard to cytokine and chemokine secretion as well as expression of cell surface antigens may instigate the generation of immune-regulatory cells. A crucial role for the presence of a microflora in the induction of regulatory T cells has been demonstrated in a murine transfer model of colitis . Protective T cells showed reduced efficacy in preventing colitis development and demonstrated CH5424802 reduced release of IL-10 and IFN-γ Acalabrutinib purchase when derived from axenic mice as opposed to those derived from conventionally housed mice. While we did not detect a significant increase in systemic T cells with a common
regulatory phenotype, such as CD25-positive T cells, we cannot exclude the generation of a specific population of cells with regulatory function in mucosal tissues and/or systemically. The increased CD11b-positive leucocyte population may be involved in the suppression of activated T cell responses. Myeloid-derived suppressor cells with a CD11b-positive, Gr-1-positive phenotype and immunosuppressive function have been described and have been implicated in SPTBN5 the protection of T cell-mediated chronic enterocolitis [26,27]. We have demonstrated previously a similar rapid onset of proinflammatory cytokine and intestinal injury in adult axenic IL-10 gene-deficient mice following colonization with commensal faecal flora . A similar uncontrolled proinflammatory cytokine response to commensal bacterial antigens has also been found to play a crucial role in the human leucocyte antigen-B27 (HLA-B27) transgenic rat
colitis model . Our results demonstrate for the first time that bacterial colonization in wild-type mice initially triggers a similar proinflammatory immune response, causing temporary intestinal inflammation. Endogenous bacterial antigens are treated as ‘foreign’ and stimulate an antigen-specific systemic immune response. However, in contrast to colitis susceptible rodents, wild-type mice are able to down-regulate the initial proinflammatory immune response and establish mucosal as well as systemic tolerance. Acquisition of immunological homeostasis appears to follow a defined inflammatory response pattern when first exposed to faecal bacteria and antigens, which probably plays an important role in the induction of tolerance to the endogenous microflora.
MAPKs are highly conserved signal transduction pathways important in the function and differentiation . In the case of DC, three specific Imatinib clinical trial pathways have been identified as important components of normal DC physiology. Stimulation of the p38 MAPK has been observed to be critical for normal maturation and function of DC . Specifically, p38 activation has been implicated in the regulation of the
surface expression of CD80, CD86, CD40, CCR7 and MHC-II molecules as well as cytoskeletal rearrangement, endocytosis, cytokine secretion and response [18–25]. Stimulation of the c-Jun N-terminal kinase (JNK) pathway has been found to be important in CD80 and CD86 expression as well as expression of CD83, MHC-II, Toll-like receptor (TLR) function, cytokine secretion and response and T cell stimulation [26–31]. Activation of the extracellular-regulated kinase (ERK) MAPK pathway has been observed contribute to TLR function and cytokine production and responsiveness [32–34]. During most viral infections, mature DC are responsible for the presentation of viral antigens to learn more naive T cells within secondary lymphoid organs, resulting in the generation of an
antigen-specific adaptive immune response and clearance of the virus . However, this is not the case with human immunodeficiency virus (HIV-1) infection . During infection with HIV-1, the virus is not cleared and a chronic systemic infection develops characterized by immune dysfunction, CD4+ T cell depletion, systemic inflammation and opportunistic infections [37–40]. How the virus evades immune system elimination is not completely understood. It has been suggested that initial HIV-1 interactions with DC may actually enhance viral spread to naive T cells in secondary lymphoid tissue. Rather than process and present critical viral antigens to induce a virus-specific adaptive immune
response, there have been reports suggesting that DC enhance HIV-1 dissemination during infection via the transfer of intact cell surface and endosomal viral particles to naive T cells in the secondary lymphoid organs [41,42]. HIV-1 itself does not appear to stimulate the maturation of DC but, rather, may induce DC dysfunction, inhibit maturation and reduce DC numbers in vivo[43–46], Thiamet G although there are reports that suggest otherwise [47–54]. In fact, a number of HIV-1-derived peptides have also been observed to induce maturation of DC [55–57]. To describe more comprehensively the effects of HIV-1 on DC, we expanded upon previous studies of the influence of HIV-1 on DC maturation and function. In addition to investigating the effects of HIV-1 infection on the expression of surface molecules pertinent to DC maturation, we studied simultaneously the effects of HIV-1 on DC function, including endocytosis, antigen presentation and cell signalling, in response to bacterial lipopolysaccharide (LPS).