Common and Distinct Clinical Features in Adult Patients with Anti-Aminoacyl-tRNA Synthetase Antibodies: Heterogeneity within the Syndrome

Objective

To identify similarities and differences in the clinical features of adult Japanese patients with individual anti-aminoacyl-tRNA synthetase antibodies (anti-ARS Abs).

Methods

This was a retrospective analysis of 166 adult Japanese patients with anti-ARS Abs detected by immunoprecipitation assays. These patients had visited Kanazawa University Hospital or collaborating medical centers from 2003 to 2009.

Results

Anti-ARS Ab specificity included anti-Jo-1 (36%), anti-EJ (23%), anti-PL-7 (18%), anti-PL-12 (11%), anti-KS (8%), and anti-OJ (5%). These anti-ARS Abs were mutually exclusive, except for one serum Ab that had both anti-PL-7 and PL-12 reactivity. Myositis was closely associated with anti-Jo-1, anti-EJ, and anti-PL-7, while interstitial lung disease (ILD) was correlated with all 6 anti-ARS Abs. Dermatomyositis (DM)-specific skin manifestations (heliotrope rash and Gottron’s sign) were frequently observed in patients with anti-Jo-1, anti-EJ, anti-PL-7, and anti-PL-12. Therefore, most clinical diagnoses were polymyositis or DM for anti-Jo-1, anti-EJ, and anti-PL-7; clinically amyopathic DM or ILD for anti-PL-12; and ILD for anti-KS and anti-OJ. Patients with anti-Jo-1, anti-EJ, and anti-PL-7 developed myositis later if they had ILD alone at the time of disease onset, and most patients with anti-ARS Abs eventually developed ILD if they did not have ILD at disease onset.

Conclusion

Patients with anti-ARS Abs are relatively homogeneous. However, the distribution and timing of myositis, ILD, and rashes differ among patients with individual anti-ARS Abs. Thus, identification of individual anti-ARS Abs is beneficial to define this rather homogeneous subset and to predict clinical outcomes within the “anti-synthetase syndrome.”     

Osmotic stress activates the TAK1-JNK pathway while blocking TAK1-mediated NF-kappaB activation: TAO2 regulates TAK1 pathways

Osmotic stress activates MAPKs, including JNK and p38, which play important roles in cellular stress responses. Transforming growth factor-beta-activated kinase 1 (TAK1) is a member of the MAPK kinase kinase (MAPKKK) family and can activate JNK and p38. TAK1 can also activate IkappaB kinase (IKK) that leads to degradation of IkappaB and subsequent NF-kappaB activation. We found that TAK1 is essential for osmotic stress-induced activation of JNK but is not an exclusive mediator of p38 activation. Furthermore, we found that although TAK1 was highly activated upon osmotic stress, it could not induce degradation of IkappaB or activation of NF-kappaB. These results suggest that TAK1 activity is somehow modulated to function specifically in osmotic stress signaling, leading to the activation of JNK but not of IKK. To elucidate the mechanism underlying this modulation, we screened for potential TAK1-binding proteins. We found that TAO2 (thousand-and-one amino acid kinase 2) associates with TAK1 and can inhibit TAK1-mediated activation of NF-kappaB but not of JNK. We observed that TAO2 can interfere with the interaction between TAK1 and IKK and thus may regulate TAK1 function. TAK1 is activated by many distinct stimuli, including cytokines and stresses, and regulation by TAO2 may be important to activate specific intracellular signaling pathways that are unique to osmotic stress.     

Papillomavirus capsid mutation to escape dendritic cell-dependent innate immunity in cervical cancer

Infection with oncogenic human papillomaviruses (HPVs), typified by HPV type 16 (HPV16), is a necessary cause of cervical cancer. Prophylactic vaccination with HPV16 L1 virus-like particles (VLPs) provides immunity. HPV16 VLPs activate dendritic cells and a potent neutralizing immunoglobulin G (IgG) response, yet many cervical cancer patients fail to generate detectable VLP-specific IgG. Therefore, we examined the role of the innate recognition of HPV16 L1 in VLP-induced immune responses and its evasion during carcinogenesis. Nonconservative mutations within HPV16 L1 have been described in isolates from cervical cancer and its precursor, high-grade cervical intraepithelial neoplasia (CIN). We determined the effect of mutations in L1 upon in vitro self-assembly into VLPs and their influence upon the induction of innate and adaptive immune responses in mice. Several nonconservative mutations in HPV16 L1 isolated from high-grade CIN or cervical carcinoma prevent self-assembly of L1 VLPs. Intact VLPs, but not assembly-defective L1, activate dendritic cells to produce proinflammatory factors, such as alpha interferon, that play a critical role in inducing adaptive immunity. Indeed, effective induction of L1-specific IgG1 and IgG2a was dependent upon intact VLP structure. Dendritic cell activation and production of virus-specific neutralizing IgG by VLPs requires MyD88-dependent signaling, although the L1 structure that initiates MyD88-mediated signaling is distinct from the neutralizing epitopes. We conclude that innate recognition of the intact L1 VLP structure via MyD88 is critical in the induction of high-titer neutralizing IgG. Tumor progression is associated with genetic instability and L1 mutants. Selection for assembly-deficient L1 mutations suggests the evasion of MyD88-dependent immune control during cervical carcinogenesis.     

Synthesis and characterization of a novel reagent containing dansyl group, which specifically alkylates sulfhydryl group: an example of application for protein chemistry

We have synthesized a novel reagent containing dansyl group, iodoacethyl dansylcadaverine (IADC), which specifically alkylates sulfhydryl groups. The carboxyl group of iodoacetic acid was activated with dicyclohexylcarbodiimide and was condensed with amino group of dansylcadaverine. Purity and chemical structure of IADC was confirmed with mass spectrometry (MS) and NMR. IADC alkylated GSH but not GSSG, which was confirmed by MS. The reactivity of IADC with proteins was also investigated with Western blotting using anti-dansyl antibody. IADC reacted only with sulfhydryl-containing proteins. The specificity of the interaction of IADC with sulfhydryl groups in proteins was confirmed by adding excessive amount of a well-known sulfhydryl-specific reagent, 5, 5'-dithiobis(2-nitrobenzoic acid), which led to a complete inhibition. To show the usefulness of IADC, the cysteines in glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from chicken muscle were modified with this reagent, and GAPDH was then digested by lysyl endopeptidase. The peptides generated from digestion of IADC-incorporated GAPDH were applied to an anti-dansyl immunoaffinity column. The peptide fragments bound and eluted from the column were separated by HPLC, and the amino acid sequence of each peptide was analyzed, and peptide was identified as the one containing a Cys residue(s). These data showed that IADC is a useful reagent to specifically identify the positions of a Cys residue(s) in proteins.     

Overt diabetes induced by overeating in neonatally STZ-treated impaired glucose tolerant mice: long-term follow up study

This study has investigated the effect of a long period of overeating on the glycemic control and pancreatic beta-cell function in neonatally streptozocin treated impaired glucose tolerant mice. Neonatally streptozocin (60 mg/kg) treated male ICR mice with 150-200 mg/dl of fed blood glucose levels were divided into two groups at 6 weeks of age. One group was maintained on a cafeteria diet (SZC) and the other on ordinary mouse chow (SZ) until 30 weeks of age. Normal male ICR mice were divided into a cafeteria diet group (CC) and an ordinary chow group (Cont). SZC and CC consumed 134-124% of the caloric intake in SZ and Cont throughout the study. Marked elevation of the fed blood glucose level was observed and the glucose tolerance was progressively impaired in SZC. On pancreas perfusion at 30 weeks of age, insulin secretion to 30 mM glucose in SZC was significantly decreased compared with that in SZ. That in CC was slightly decreased compared with that in Cont. The pancreatic insulin concentration in SZC was significantly less than that in SZ. We conclude that chronic hyperglycemia, induced by the long period of overeating, accelerated the selective loss of beta-cell sensitivity to glucose. Even in normal mice that did not have marked hyperglycemia, insulin secretion to glucose was suppressed, probably by chronic stimulation of the beta-cell due to the long period of dietary excess.     

Akt contributes to activation of the TRIF-dependent signaling pathways of TLRs by interacting with TANK-binding kinase 1

Toll/IL-1R domain-containing adaptor inducing IFN-β (TRIF) is an adaptor molecule that is recruited to TLR3 and -4 upon agonist stimulation and triggers activation of IFN regulatory factor 3 (IRF3) and expression of type 1 IFNs, which are critical for cellular antiviral responses. We show that Akt is a downstream molecule of TRIF/TANK-binding kinase 1 (TBK1) and plays an important role in the activation of IRF3 by TLR3 and -4 agonists. Blockade of Akt by a dominant-negative mutant or by short interfering RNA decreased IRF3 activation and IFN-β expression induced by polyinosinic:polycytidylic acid [poly(I:C)], LPS, TRIF, and TBK1. Association of endogenous TBK1 and Akt was observed in macrophages when stimulated with poly(I:C) and LPS. In vitro kinase assays combined with reversed-phase liquid chromatography mass spectrometry analysis showed that TBK1 enhanced phosphorylation of Akt on Ser(473), whereas knockdown of TBK1 expression by short interfering RNA in macrophages decreased poly(I:C)- and LPS-induced Akt phosphorylation. Embryonic fibroblasts derived from TBK1 knockout mice also showed impaired Akt phosphorylation in response to poly(I:C) and LPS. To our knowledge, our results demonstrate a new regulatory mechanism for Akt activation mediated by TBK1 and a novel role of Akt in TLR-mediated immune responses.     

Wakubitinema toyamai n. gen. and n. sp. (Nematoda: Seuratoidea: Quimperiidae) from the intestine of Rana (Limnonectes) Namiyei (Amphibia: Ranidae) on Okinawa Island, Japan

Wakubitinema n. gen. (Nematoda: Seuratoidea: Quimperiidae: Quimperiinae) is erected for Wakubitinema toyamai n. sp. from the small intestine of Rana (Limnonectes) namiyei Stejneger, 1901, on Okinawa Island, Japan. Wakubitinema resembles Paraquimperia Baylis, 1934, and Desmognathinema Baker et al., 1987, but is readily distinguished from the former genus by the distinctly divided esophagus and the absence of cervical flanges and lateral alae, and from the latter genus by the postesophageal position of the excretory pore and cervical papillae and the presence of preanal unpaired papilla in males. Close morphological similarities between Wakubitinema and Paraquimperia may suggest that Wakubitinema has evolved from a quimperiid fish.     

A Sialylated Voltage-Dependent Ca2+ Channel Binds Hemagglutinin and Mediates Influenza A Virus Entry into Mammalian Cells

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Induction and characterization of minor histocompatibility antigens. Specific primary cytotoxic T lymphocyte responses in vitro

A definite cytotoxic activity was developed in a BALB/c (H-2d) anti-DBA/2 primary mixed leukocyte culture (MLC), which received interleukin 2 (IL-2) on day 3 of culture. This cytotoxic activity was minor histocompatibility antigens (MIHA)-specific at the stimulator level, and was not developed in a syngeneic (BALB/c anti-BALB/c) MLC. The addition of IL-2 on day 3 of culture was crucial; no or very weak cytotoxic activity was developed in MLC receiving IL-2 on day 0 or on both day 0 and day 3. Only appropriate MIHA-allogeneic tumor cells were lysed as the target of the cytotoxic activity. The cytotoxic activity seemed MIHA-specific also at the target level; it lysed tumor cells of DBA/2 mouse origin but not those of BALB/c (syngeneic) origin. Phenotypes of the cytotoxic effector cell were Thy-1+ Lyt-2+. We concluded from these results that MIHA-specific cytotoxic T lymphocytes (CTL) were generated in the MIHA-allogeneic primary MLC. In this newly developed system, we studied genetic and antigenic requirements for primary anti-MIHA CTL responses in vitro. We demonstrated; among spleen cells (SC) of seven B10 H-2-congenic strains only SC of B10.D2 strain whose major histocompatibility complex (MHC) (H-2d) was compatible with the responder MHC effectively stimulated responder BALB/c (H-2d) SC for an anti-MIHA (DBA-C57BL-common) CTL response. Similarly, only SC of two out of seven C x B recombinant inbred strains (C x B.H and C x B.D), which were compatible at the MHC with responder SC, activated responder BALB/c SC for the response. The possibility that cells responding to H-2 alloantigens suppressed the anti-MIHA response was ruled out. Additional experiments showed that compatibility at the H-2K-end or the H-2D-end of the MHC was sufficient for a definite anti-MIHA response. These provided formal evidence that primary anti-MIHA CTL responses in vitro were MHC-restricted at the stimulator level. We then showed that sonication-disrupted SC or Sephadex G-10 column-passed nonadherent SC failed to stimulate responder SC for a primary anti-MIHA CTL response, whereas G-10-passed nonadherent SC responded well to adherent stimulator cells. Further study demonstrated that Ia+ adherent cells were the most active cell type as stimulator. Finally, we confirmed that the primary anti-MIHA CTL responses to adherent stimulator cells was MHC-restricted.