The patients, ranging in age from 21 to 78 years (mean, 51.3 years) Ganetespib ic50 and having adequate liver function reserve, had survived for at least 2 months after hepatectomy, and none received treatment prior to surgery such as transarterial chemoembolization or radiofrequency ablation. Clinicopathologic features of the 120 HCCs in this study are described in Table 1. Surgically resected specimens were partly embedded in paraffin after fixation in 10% formalin for histological processing and

partly immediately frozen in liquid nitrogen and stored at -80°C. All available hematoxylin and eosin stained slides were reviewed. The tumor grading was based on the criteria proposed by Edmondson and Steiner (I, well differentiated; II, moderately differentiated; III, poorly

differentiated; IV, undifferentiated) [16]. The conventional TNM system outlined in the cancer staging manual (6th ed.) by the American Joint Committee on Cancer (AJCC) was used in tumor staging. Table 1 Relations between NNMT mRNA levels and clinicopathologic features in HCC   All patients (n = 120)   Clinicopathologic parameters High NNMT (n = 48) Copy number ratio ≥ 4.40 Low NNMT (n = 72) Copy number ratio < 4.40 P value Age     0.730 < 55 years 31 43   ≥ 55 years 17 29   Gender     0.758 Male 38 54   Female 10 Palbociclib mouse 18   HbsAg     0.885 Absent 8 14   Present 40 58   HCV     0.823 Absent 45 67   Present 3 5   Liver cirrhosis     0.852 Absent 25 40   Present 23 32   Tumor stage     0.010 I 23 23   II 9 33   III & IV 16 16   AFP level     0.314 < 100 ng/ml 28 34   ≥ 100 ng/ml 20 38   Tumor size     0.733 < 5 cm 27 44   ≥ 5 cm 21 28   Edmondson grade     0.368 I 13 15   II 30 43   III & IV 5 mafosfamide 14   RNA extraction and cDNA synthesis Total RNA was extracted from cancerous and surrounding non-cancerous frozen tissues using an RNeasy minikit (Qiagen, Germany) according to the manufacturer’s instructions. The integrity

of all tested total RNA samples was verified using a Bioanalyzer 2100 (Agilent Technologies, United States). DNase I treatment was routinely included in the extraction step. Residual genomic DNA contamination was assayed by a quantitative real-time PCR assay for GAPDH DNA and samples with contaminating DNA were re-subjected to DNase I treatment and assayed again. Samples containing 4 μg of total RNA were incubated with 2 μl of 1 μM oligo d(T)18 primer (Genotech, Korea) at 70°C for 7 min and cooled on ice for 5 min. The enzyme mix was separately prepared in a total volume of 11 μl by adding 2 μl of 0.1 M DTT (Duchefa, Netherlands), 2 μl of 10× reverse-transcription buffer, 5 μl of 2 mM dNTP, 1 μl of 200 U/μl MMLV reverse-transcriptase, and 1 μl of 40 U/μl RNase inhibitor (Enzynomics, Korea). After adding the enzyme mix to the annealed total RNA sample, the reaction was incubated for 90 min at 42°C prior to heat inactivation of reverse-transcriptase at 80°C for 10 min.

Cancer 2005,104(10):2099–2103.PubMed 227. Barkholt L, Bregni M, Remberger M, Blaise D, Peccatori J, Massenkeil G, Pedrazzoli P, Zambelli A, Bay JO, Francois S, et al.: Allogeneic

haematopoietic stem cell transplantation for metastatic renal carcinoma in Europe. Ann Oncol 2006,17(7):1134–1140.PubMed 228. Artz AS, Van Besien K, Zimmerman T, Gajewski TF, Rini BI, Hu HS, Stadler WM, Vogelzang NJ: Long-term follow-up of nonmyeloablative allogeneic stem cell transplantation for renal cell carcinoma: The University of Chicago Experience. Bone Marrow Transplant 2005,35(3):253–260.PubMed 229. Childs R, Chernoff A, Contentin N, Bahceci E, Schrump BGB324 D, Leitman S, Read EJ, Tisdale J, Dunbar C, Linehan WM, et al.: Regression of metastatic renal-cell carcinoma after nonmyeloablative allogeneic peripheral-blood stem-cell transplantation. N Engl J Med 2000,343(11):750–758.PubMed 230. Singletary SE: Breast cancer management: the road to today. Cancer 2008,113(7 Suppl):1844–1849.PubMed 231. Biron P, Durand M, Roche H, Delozier T, Battista C, Fargeot P, Spaeth D, Bachelot T, Poiget E, Monnot

F, et al.: Pegase 03: a prospective randomized phase III trial of FEC with or without high-dose thiotepa, cyclophosphamide and autologous stem cell transplantation in first-line treatment of metastatic breast cancer. Bone Marrow this website Transplant 2008,41(6):555–562.PubMed 232. Ueno NT, Rizzo JD, Demirer T, Cheng YC, Hegenbart U, Zhang MJ, Bregni M, Carella A, Blaise D, Bashey A, et al.: Allogeneic hematopoietic cell

transplantation for metastatic breast cancer. Bone Marrow Transplant 2008,41(6):537–545.PubMed 233. Carella AM, Bregni M: Current role of allogeneic stem cell transplantation in breast cancer. Ann Oncol 2007,18(10):1591–1593.PubMed 234. Gill S, Blackstock AW, Goldberg RM: Colorectal cancer. Mayo Clin Proc 2007,82(1):114–129.PubMed 235. Benson AB: Epidemiology, disease PLEKHB2 progression, and economic burden of colorectal cancer. J Manag Care Pharm 2007,13(6 Suppl C):S5–18.PubMed 236. Nagy VM: Updating the management of rectal cancer. J Gastrointestin Liver Dis 2008,17(1):69–74.PubMed 237. Kojima R, Kami M, Hori A, Murashige N, Ohnishi M, Kim SW, Hamaki T, Kishi Y, Tsutsumi Y, Masauzi N, et al.: Reduced-intensity allogeneic hematopoietic stem-cell transplantation as an immunotherapy for metastatic colorectal cancer. Transplantation 2004,78(12):1740–1746.PubMed 238. Aglietta M, Barkholt L, Schianca FC, Caravelli D, Omazic B, Minotto C, Leone F, Hentschke P, Bertoldero G, Capaldi A, et al.: Reduced-intensity allogeneic hematopoietic stem cell transplantation in metastatic colorectal cancer as a novel adoptive cell therapy approach. The European group for blood and marrow transplantation experience. Biol Blood Marrow Transplant 2009,15(3):326–335.PubMed 239. Hashino S, Kobayashi S, Takahata M, Onozawa M, Nakagawa M, Kawamura T, Fujisawa F, Izumiyama K, Kahata K, Kondo T, et al.

20 μm pore size filter and frozen in 40 ml aliquots. Immediately prior to use, the sterile saliva was thawed at 37°C; the slight precipitate was pelleted at 1,430 × g for 5 min, and the clear Erlotinib ic50 25% saliva supernatant was used in experiments. Microscope observation Quantitative and structural analysis of homotypic P. gingivalis biofilms was accomplished by confocal laser scanning microscopy (CLSM, Radiance 2100, Bio-Rad) and subsequent image

analysis [50]. P. gingivalis was stained with CFSE (8 μg/ml; Molecular Probes, Eugene, OR), washed three times and 1 × 108 cells in PBS or dTSB were anaerobically incubated in a 25% saliva-coated wells of a chambered coverglass system (Culture Well™, Grace Bio Labs, Bend, OR) for 24 hours at 37°C in the dark on a rotator. The resulting biofilms were examined using the CLSM with reflected laser light at 488 nm. The images were analyzed using the Image J 1.34s (National Institutes of Health; Bethesda, MD) and Imaris 5.0.1 (Bitplane AG; Zurich, Switzerland) software packages. The experiment was repeated independently three times with each strain in triplicate. Biofilm characterization by image

analysis Z stacks of the x-y sections find more in the CLSM images were converted to composite images with the “”Iso Surface”" function of the “”Surpass”" option provided by Imaris 5.0.1 (Bitplane AG; Zurich, Switzerland) software. Iso Surface images were created at a threshold of 40 and smoothed with the “”Gaussian Filter”" function at a width of 1.28 μm, then the biovolume was calculated. Measurement of peak parameters was performed as described previously [50]. Digitally reconstructed images of the x-z section,

189.4 μm × appropriate height with 10-μm spaced y-series slices, were created using the “”Reslice”" function of Image J. An image series of the x-z section was processed using the “”Find Edges”" Teicoplanin function, then the peak height was calculated by Image J. Color images of the x-z section were converted into gray scale and the density per vertical position (x-axis) was analyzed with the “”Plot profile”" function of Image J. The data were then exported as plot values with x-axis distance information. Peaks were defined as positions where plot values were higher than on either side, and the distance between two peaks was measured. The peak number was counted in a 90-μm section of the x-axis. Exopolysaccharide production assay P. gingivalis organisms were stained with DAPI (50 μg/ml; Molecular Probes, Eugene, OR), then washed and cultured in 25% saliva-coated wells of CultureWell chambered coverglass system with dTSB for 24 hours. The resulting biofilms were washed, then exopolysaccharide was labelled with Concanavalin A-FITC and Wheat germ agglutinin-FITC (100 μg/ml; Molecular Probes) for 30 minutes at room temperature, as described previously [10]. After washing, fluorescent images were obtained using CLSM with reflected laser light at 405 and 488 nm, then analyzed as described above.

As a proliferation inhibitor, p21Waf1/cip1 was chosen because it is poised to play an important role in preventing tumor development. Cyclin D1-CDK4 complexes promote G1 selleck chemicals llc phase progression through phosphorylation and inactivation of the retinoblastoma (Rb) gene product. Our results showed that specific downregulation of STIM1 inhibited human glioblastoma cell proliferation and induced G0/G1 phase cell cycle arrest by increasing expression of p21Waf1/cip1 and decreasing expression of Cyclin D1-CDK4. Therefore, STIM1 may serve as a therapeutic target for human glioblastoma. Methods Reagents and antibodies Dulbecco’s modified Eagle’s medium (DMEM),

fetal bovine serum (FBS), TRIzol® Reagent and Lipofectamine™ 2000 were purchased from Invitrogen (Carlsbad, CA); 3-(4,5-dimethylthylthiazol-2yl-)-2,5-diphenyl tetrazolium bromide (MTT) RAD001 cost (Dingguo biology, Shanghai, China); Dimethylsulfoxide (DMSO) (Shanghai Sibas Biotechnology Development Co., Ltd., China); 5-Bromo-2-deoxyuridine (BrdU) Cell Proliferation ELISA kit was purchased from Roche Applied Sciences (Indianapolis, IN); Giemsa was purchased from Chemicon International (Temecula, CA); Propidium Iodide (PI) was purchased from Sigma-Aldrich (St. Louis, MO); Bicinchoninic

acid (BCA) Protein assay was purchased from HyClone-Pierce (South Logan, UT); M-MLV Reverse Transcription was purchased from Promega (Madison, WI); Oligo-dT was purchased from Sangon Biotech (Shanghai, China); SYBR green Master Mixture was purchased from Takara (Otsu, Japan); pFH-L vector and virion-packaging elements (packing plasmid mix) were obtained from Holybiol (Shanghai, China). Mouse anti-STIM1, mouse anti-GAPDH, p21Waf1/Cip1 , cyclin D1, cyclin-dependent kinase 4 (CDK4) and goat anti-mouse IgG were purchased from Santa Cruz biotechnology (Santa Cruz, CA), mouse anti-STIM2 was purchased from Abcam plc (Abcam, UK), mouse anti-Orai1 was purchased from Sigma biotechnology (Sigma- -Aldrich, US). All other

chemicals were of analytical grade. Cell culture Human kidney cell line HEK293,human glioblastoma cell lines, U251, U87 and U373, SPTLC1 were all obtained from the American Type Culture Collection (ATCC, Manassas, VA) and cultured in DMEM containing 10% FBS, 100U/mL penicillin and 100 μg/mL streptomycin at 37°C in a humidified atmosphere containing 5% CO2. siRNA design and construction of recombinant lentiviral vector Recombinant lentiviral vector was constructed as described previously [19]. The candidate sequence (5′-CCTGGATGATGTAGATCATAA-3′) in the STIM1 cDNA sequence (GenBank accession number NM_003156) was selected for siRNA and blasted against the human genome database to eliminate cross-silence phenomenon with non-target genes. Scrambled siRNA (5′-TTCTCCGAACGTGTCACGT-3′) that does not target any genes was used as the negative control.

Major histocompatibility complex (MHC) class-I H-2kd-restricted cognate antigenic peptides islet-specific glucose-6-phosphatase

catalytic subunit-related protein (IGRP206–214) (VYLKTNVFL) and its mimotopes NRP (KYNKANWFL; agonist), NRP-V7 (KYNKANVFL; super agonist) and TUM (KYQAVTTTL; non-agonist) PR-171 cost were custom synthesized by Genscript (Piscataway, NJ, USA). Expression of cell surface markers was evaluated by flow cytometry using fluorescence activated cell sorter (FACS)Canto flow cytometer (Becton Dickinson Flow Cytometry Systems, San Jose, CA, USA) and the data were analysed using FlowJo software (Tree Star Inc., Ashland, OR, USA). Total lymph node cells (2 × 105 cells) or purified CD8+ T cells (2·5 × 104 cells) were cultured in 96-well culture plates with the indicated peptides using irradiated splenocytes as antigen-presenting cells (APCs)

(1 × 105 cells) or with anti-CD3/CD28-coated beads for 72 h. Cell proliferation was measured by [3H]-thymidine incorporation [34]. To measure antigen-induced proliferation in vivo, 8.3 CD8+ T cells were labelled with AZD9668 chemical structure carboxyfluorescein diacetate succinimidyl ester (CFSE), as described previously [35], and injected intravenously. Bone marrow-derived dendritic cells (BMDCs) cultured with granulocyte–macrophage colony-stimulating factor (GM-CSF) and IL-4 were pulsed with IGRP206–214 or the control peptide TUM for 1 h at 37°C, washed, resuspended in phosphate-buffered saline (PBS) and injected

subcutaneously in hind footpads. Donor cells recovered from the draining inguinal lymph node were evaluated to measure proliferation. CTL activity was measured using RMA-S-Kd target cells loaded with the cognate peptide, as described previously [1, 32]. The amount of IL-2 in the culture supernatants was determined by sandwich ELISA using antibody pairs purchased from BD Pharmingen Biosciences (Palo Alto, CA, USA). Onset of T1D was monitored by measuring urine glucose levels using Keto-Diastix (Bayer, Canada). ADP ribosylation factor Animals with two consecutive readings of >3 were considered diabetic. At the time of euthanasia, pancreatic tissues were processed for histopathology analysis. At least three non-overlapping (200 μm apart) 5-μm sections were evaluated for insulitis [32]. Cumulative incidence of T1D was analysed using Prism software (GraphPad Software Inc., La Jolla, CA, USA). For diabetes incidence, significance was calculated using log-rank (Mantel–Cox) test. For all other parameters, statistical significance was calculated by Student’s t-test. The 8.3-NOD mouse expresses a highly pathogenic, MHC class I-restricted, transgenic 8.3 TCR specific to a peptide derived from the IGRP206–214 [33, 36]. In these mice, the 8.3 TCR transgenic CD8+ T cells (8.3 T cells) infiltrate pancreatic islets from 3 weeks of age [33]. Female 8.

However, MHC class I molecules often also contain a number of unpaired cysteine residues, most notably at position 67 in the peptide-groove, which in the case of HLA-B27 has been shown to be involved in the formation of partially unfolded heavy-chain homodimers,8–10 and at position HDAC inhibition 42 on the

external face of the molecule, which in HLA-G allows the formation of fully folded dimers.11,12 Significantly, there are also unpaired cysteine residues in the transmembrane domain region of HLA-B molecules at position 308, and in the cytoplasmic tail domain of many HLA-B molecules at position 325, and at position 339 in HLA-A molecules. DAPT in vitro The precise role, if any, of these cysteine residues remains unclear, though modification by palmitylation,7 involvement in dimer formation,13 transient interactions in the MHC class I peptide-loading complex,14 and NK receptor recognition have all been demonstrated.7 We recently identified that the cytoplasmic tail domain cysteines were intimately involved in the formation of fully folded MHC class I dimers in exosomes.15 These 50–150 nm vesicles form in the endocytic pathway in multivesicular bodies, some of which are released into the extracellular environment.16 They are released by a wide range

of both normal and tumour cells, and have been implicated in a number of biological processes. We established that the formation of MHC class I dimers in exosomes

was a function of the low level of glutathione (GSH) detected in these vesicles when compared with whole cell lysates, and hypothesized that exosomes cannot maintain the reducing Reverse transcriptase environment of the normal cytoplasm, hence allowing disulphide bonds to form between the cytoplasmic tails.15 To address whether there were also circumstances wherein MHC class I dimers could be induced to form by mimicking the low GSH levels seen in exosomes, we set up experimental systems to modify the cellular redox environment, both by using a strong oxidant treatment, and by inducing apoptosis with agents known to cause a depletion of intracellular GSH. Our data indicate that apoptosis-induced alterations to cellular redox do indeed lead to the induction of MHC class I dimers. The human lymphoblastoid lines .221 (gifted by Salim Khakoo, Imperial College, London, UK) and CEM (gifted by Antony Antoniou, UCL, London, UK), the human Epstein–Barr virus-transformed B-cell line Jesthom (Health Protection Agency line no. 88052004), and the rat C58 thymoma line (gifted by Geoff Butcher; Babraham Institute, Cambridge, UK) were cultured in RPMI-1640 (Gibco, Paisley, UK) supplemented with 10% fetal bovine serum (Gibco).

7,25 The recognition

of cells by the immune system when undergoing redox stress is not well Selleck Navitoclax defined. Our data suggest that cells undergoing a redox stress that leads to a lowering in the levels of intracellular glutathione may begin to display MHC class I dimers on their cell surface. Such alterations in cellular glutathione levels have been reported to occur during T-cell activation26,27 and also during apoptosis.17,18,28 A more extensive study monitoring both MHC class I dimer formation and the level of glutathione in cells undergoing a variety of stimuli would, we consider, be of some worth. Furthermore, distinguishing whether both apoptotic and necrotic cell death pathways induce dimers would also be informative, alongside other pathological states such as viral infection, Ruxolitinib molecular weight and many others that induce inflammatory responses and the production of reactive oxygen species. This would allow a pattern of conditions to be catalogued under which MHC class I dimer formation is induced. Although various MHC class I dimers have been reported in the literature, their potential biological role remains enigmatic.8,13 Of significant note, however, are the observations that Ig-MHC class I dimers can act to tolerize T cells.29 In this study, Kourilsky and colleagues used a soluble H2-Kd molecule dimerized with a cross-linking antibody to demonstrate

that an antigen-specific T-cell hybridoma was initially activated, followed by a state of unresponsiveness. It has also been demonstrated that antigen-specific occupancy of just one of the two peptide-binding grooves in an

Coproporphyrinogen III oxidase MHC class I dimer can have an effect on T cells,30 which may allow not only the HLA-B dimers we show here, but also the HLA-A-B dimers we describe in Fig. 2 to have some biological activity. Hence, the MHC class I dimers detected on apoptotic cells, and also on exosomes,15 may be capable of providing signal, including tolerogenic signals to immune cells. Similarly, tumour cells undergoing apoptosis, or releasing exosomes containing tumour-associated or tumour-specific antigens may influence T-cell behaviour. Of further interest is the possible recognition of MHC class I dimers by members of the NK cell receptor family. It has been shown that a disulphide-linked engineered version of the KIR2DL1 receptor has an increased affinity for HLA-C,31 and that KIR molecules can form an array of dimers and multimers in a zinc-dependent interaction.32 Hence interactions between dimers of both ligands and receptors may occur, potentially inducing extra stability for the generation of either inhibitory or activatory signals. As indicated above, defining the various conditions under which such dimers form would allow the design of experiments to directly study their potential influence on immune responses.

64±10.87×106 and WT: 31.54±15.52×106 for B220+; Hax1−/−: 3.71±0.77×106 and WT: 2.55±1.05×106 for T1; Hax1−/−: 6.91±3.61×106 and WT: 4.73±2.23×106 for T2; Hax1−/−: 5.89±2.89×106 and WT: 4.53±2.39×106 for mature B cells; Hax1−/−: 2.92±1.84×106 and WT: 2.34±1.16×106 for MZ B cells). Our data clearly demonstrate that Hax1−/− LSK cells in a Hax1+/+ environment were able to fully reconstitute the lethally irradiated hosts. To further investigate the reason for the massive B lymphocyte deficiency, we investigated Selleckchem Paclitaxel the expression of CXCR4 and BAFFR on splenic B cells. CXCR4 is expressed on hematopoietic precursors 22 as well as on centroblasts within the germinal centre

18. CXCR4-expressing cells migrate towards CXCL12, expressed by stromal cells and germinal center dark zone compartments. Thus, an impaired CXCR4 expression would severely impede normal B-cell development. Alternatively, signals through the BAFFR have a significant role in promoting B-cell survival and homeostatic proliferation 23. For real time analysis, we isolated total splenocytes of four 10-wk-old WT and Hax1−/− mice and enriched for B lymphocytes using magnetic cell sorting. Both the CXCR4 and the BAFFR buy PLX4032 amplification showed prominent amplification products. Most interestingly, CXCR4 expression

in HAX1-deficient B cells was decreased by around 70% compared to WT cells. BAFFR expression was slightly, but not significantly, decreased in HAX1-deficient B cells (Fig. 7A). However, the decreased expression had no effect on the formation of follicular structures. No differences in the distribution of B- Rutecarpine and T-cell areas, as stained by CD3 and B220, were detectable (Fig. 7B). Because of the fact that the transfer of Hax1−/− bone marrow cells into a HAX1+ environment gave rise to normal levels of B220+ cells and functional B-cell subsets, we conclude that the severely decreased

CXCR4 expression on HAX1-deficient B cells is not solely responsible for the described B-cell loss in Hax−/− mice. Previously, we described HAX1 as interaction partner of membrane bound IgE (mIgE) 24. From that point of view, it would have been of most interest to analyse IgE responses on a Hax1-deficient background. However, the short lifespan of Hax1−/− mice impeded a direct analysis. Therefore, we focused on the detailed investigation of the biological function of HAX1 during lymphocyte development. Hax1−/− mice are characterized by a severely diminished cellularity of lymphoid tissues accompanied by a significant reduction of B and Tlymphocytes. Recently, Chao et al. 25 reported on the role of HAX1 with a similar approach. Our results demonstrate that the developmental impairment is not restricted to specific developmental stages. We observed reduced numbers of B cells from the pro-pre B-cell stage in the bone marrow to mature stages in the spleen. The analysis of splenic subpopulations clearly demonstrated a continuation of the developmental defects for T1 and T2 B cells 26, 27.

These

results suggest that pyriproxyfen is a safe chemical. Moreover, unlike alum, pyriproxyfen induces an increase in titers of IgG2a check details and enhanced TNF-α and IFN-γ. These observations indicate that the mechanism of immune enhancement by pyriproxyfen may differ from that which has been well established for alum. The authors are grateful to the students of the Department of Microbiology, Faculty of Pharmaceutical Sciences, Fukuoka University, for their cooperation during these experiments. The first author was supported by a scholarship from the Ministry of Science and Education, Japan. None of the authors has any conflict of interest associated with this study. “
“M3 muscarinic acetylcholine receptor (M3R) plays a crucial role in the secretion of saliva from salivary glands. It is reported that some patients with Sjögren’s syndrome (SS) carried inhibitory autoantibodies against M3R. The purpose of this study is to clarify the epitopes and function of anti-M3R antibodies in SS. We synthesized peptides encoding the extracellular domains of human-M3R including the N-terminal region and the

first, second and third extracellular loops. Antibodies against these regions were examined by enzyme-linked immunosorbent assay in sera from 42 SS and 42 healthy controls. For functional analysis, human salivary gland (HSG) cells were preincubated with immunoglobulin G (IgG) separated from sera of anti-M3R antibody-positive SS, -negative SS and controls for 12 h. After loading

with Fluo-3, HSG cells were stimulated with cevimeline hydrochloride, selleck compound library and intracellular Ca2+ concentrations [(Ca2+)i] were measured. Antibodies to the N-terminal, first, second and third loops were mafosfamide detected in 42·9% (18 of 42), 47·6% (20 of 42), 54·8% (23 of 42) and 45·2% (19 of 42) of SS, while in 4·8% (two of 42), 7·1% (three of 42), 2·4% (one of 42) and 2·4% (one of 42) of controls, respectively. Antibodies to the second loop positive SS-IgG inhibited the increase of (Ca2+)i induced by cevimeline hydrochloride. Antibodies to the N-terminal positive SS-IgG and antibodies to the first loop positive SS-IgG enhanced it, while antibodies to the third loop positive SS-IgG showed no effect on (Ca2+)i as well as anti-M3R antibody-negative SS-IgG. Our results indicated the presence of several B cell epitopes on M3R in SS. The influence of anti-M3R antibodies on salivary secretion might differ based on these epitopes. Sjögren’s syndrome (SS) is an autoimmune disease that affects exocrine glands, including salivary and lacrimal glands. It is characterized by lymphocytic infiltration into exocrine glands, leading to dry mouth and eyes. A number of autoantibodies, such as anti-SS-A and SS-B antibodies, are detected in patients with SS. However, no SS-specific pathological autoantibodies have yet been found in this condition [1].