Aryl hydrocarbon receptor-targeted therapy for CD4+ T cell-mediated idiopathic pneumonia syndrome in mice.

We previously demonstrated that interferon γ (IFN-γ) derived from donor T cells co-opts the indoleamine 2,3-dioxygenase 1 (IDO1) → aryl hydrocarbon receptor (AHR) axis to suppress idiopathic pneumonia syndrome (IPS). Here we report that the dysregulated expression of AP-1 family genes in Ahr-/- lung epithelial cells exacerbated IPS in allogeneic bone marrow transplantation settings. AHR repressed transcription of Jund by preventing STAT1 from binding to its promoter. As a consequence, decreased interleukin-6 impaired the differentiation of CD4+ T cells toward Th17 cells. IFN-γ- and IDO1-independent induction of Ahr expression indicated that the AHR agonist might be a better therapeutic target for IPS than the IDO1 activator. We developed a novel synthetic AHR agonist (referred to here as PB502) that potently inhibits Jund expression. PB502 was highly effective at inducing AHR activation and ameliorating IPS. Notably, PB502 was by far superior to the endogenous AHR ligand, L-kynurenine, in promoting the differentiation of both mouse and human FoxP3+ regulatory CD4+ T cells. Our results suggest that the IDO1-AHR axis in lung epithelial cells is associated with IPS repression. A specific AHR agonist may exhibit therapeutic activity against inflammatory and autoimmune diseases by promoting regulatory T-cell differentiation.

Priming Mesenchymal Stem/Stromal Cells with a Combination of a Low Dose of IFN-γ and Bortezomib Results in Potent Suppression of Pathogenic Th17 Immunity Through the IDO1-AHR Axis.

Preconditioning of mesenchymal stem/stromal cells (MSCs) with the inflammatory cytokine IFN-γ enhances not only their immunosuppressive activity but also their expression of HLA and proinflammatory genes. We hypothesized that prevention of the upregulation of inflammatory cytokines and HLA molecules in IFN-γ-primed MSCs would render these cells more immunosuppressive and less immunogenic. In this study, we discovered the following findings supporting this hypothesis: (1) activated human T cells induced the expression of IDO1 in MSCs via IFN-γ secretion and those MSCs in turn inhibited T-cell proliferation in an AHR-dependent fashion; (2) there was no difference in the expression of IDO1 and HLA-DR in MSCs after priming with a low dose (25 IU/mL) versus a high dose (100 IU/mL) of IFN-γ; (3) the transient addition of bortezomib, a proteasome inhibitor, to culture MSCs after IFN-γ priming decreased the expression of HLA-DR, inflammatory cytokine genes and Vcam1 while increasing the expression of IDO1 and the production of L-kynurenine; finally, MSCs primed with a combination of a low dose of IFN-γ and bortezomib were more effective in inhibiting Th17-mediated idiopathic pneumonia syndrome (IPS) and chronic colitis than unprimed MSCs. Our results suggest that bortezomib significantly eliminates the unfavorable effects of IFN-γ priming of MSCs (increased expression of MHC molecules and inflammatory cytokines and cell aggregation genes) and simultaneously increases their immunosuppressive activity by upregulating IDO1. Taken together, our newly established MSC priming method may contribute to MSC-based cell therapy for inflammatory diseases.

Influence of Chemical and Genetic Manipulations on Cellular Organelles Quantified by Label-Free Optical Diffraction Tomography.

Label-free optical diffraction tomography provides three-dimensional imaging of cells and organelles, along with their refractive index (RI) and volume. These physical parameters are valuable for quantitative and accurate analysis of the subcellular microenvironment and its connections to intracellular biological properties. In biological and biochemical cell analysis, various invasive cell manipulations are used, such as temperature change, chemical fixation, live cell staining with fluorescent dye, and gene overexpression of exogenous proteins. However, it is not fully understood how these various manipulations affect the physicochemical properties of different organelles. In this study, we investigated the impact of these manipulations on the cellular properties of single HeLa cells. We found that after cell fixation and an increase in temperature, the RI value of organelles, such as the nucleus and cytoplasm, significantly decreased overall. Interestingly, unlike the cell nuclei, cytoplasmic RI values were hardly detected after membrane permeation, indicating that only intracytoplasmic components were largely lost. Additionally, our findings revealed that the expression of GFP and GFP-tagged proteins significantly increased the RI values of organelles in living cells compared to the less effective RI changes observed with chemical fluorescence staining for cell organelles. The result demonstrates that distinct types of invasive manipulations can alter the microenvironment of organelles in different ways. Our study sheds new light on how chemical and genetic manipulations affect organelles.

TRAF2 regulates the protein stability of HIPK2.

A nuclear serine/threonine kinase homeodomain-interacting protein kinase 2 (HIPK2) is a critical regulator of development and DNA damage response. HIPK2 can induce apoptosis under cellular stress conditions and thus its protein level is maintained low by constant proteasomal degradation. In the present study, we present evidence that TNF receptor-associated factor 2 (TRAF2) regulates the protein stability of HIPK2. Overexpression of TRAF2 decreased while its knockdown increased the HIPK2 protein level. The TRAF2-mediated decrease in HIPK2 protein expression was blocked by proteasomal inhibitor. In addition, TRAF2 decreased the protein half-life of HIPK2. We found that HIPK2 and TRAF2 co-immunoprecipitated. Interestingly, the co-immunoprecipitation was reduced while HIPK2 protein level increased following TNFα treatment, suggesting TNFα induced dissociation of TRAF2 from HIPK2 to accumulate HIPK2. Inhibition of HIPK2 partially suppressed TNFα-induced cell death, indicating that the accumulated HIPK2 may contribute to the TNFα-induced cell death. Our results suggest that TRAF2 can regulate proapoptotic function of HIPK2 by promoting proteasomal degradation.

Characterization of Latilactobacillus curvatus MS2 isolated from Korean traditional fermented seafood and cholesterol reduction effect as synbiotics with isomalto-oligosaccharide in BALB/c mice.

This study investigated the properties of Latilactobacillus curvatus MS2 isolated from Korean traditional fermented seafood as probiotics and the effect of reducing cholesterol as a synbiotic with isomalto-oligosaccharide (IMO) in BALB/c mice. The isolated strain showed high resistance to acids and bile acids and exhibited a high DPPH scavenging capacity of 72.27 ± 0.38 %. In the intestinal adhesion test using HT-29 cells, the adhesion rate of MS2 was 17.10 ± 1.78 %, which was higher than the adhesion rate of the other investigated probiotics. MS2 showed good antimicrobial activity against food-borne pathogens, especially Staphylococcus aureus, S. epidermidis, Escherichia coli, and Vibrio vulnificus. This strain had high availability for IMO among the prebiotics of fructo-oligosaccharide, inulin and IMO. Oral administration of MS2 and IMO to BALB/c mice for 5 weeks resulted in a significant reduction in blood cholesterol levels by regulating liver lipid metabolism. These results suggest that the combination of MS2 and IMO has potential for application in functional foods.

Bilateral supernumerary clavicular heads of sternocleidomastoid muscle in a Korean female cadaver.

Many anatomical variants on the sternocleidomastoid muscle have been reported. In this study, supernumerary clavicular heads of sternocleidomastoid muscle in a Korean female cadaver were bilaterally displayed. The observed supernumerary heads were classified as follows: one sterno-mastoid, one cleido-occipital and one cleido-mastoid on the right side, and one sterno-mastoid-occipital, four cleido-occipitals, and one cleido-mastoid on the left side. The sterno-mastoid and sterno-mastoid-occipital and the cleido-occipital made the superficial layer of the sternocleidomastoid muscle, while others made deep layer. We discussed clinical relevance and developmental basis of these muscular variations important for clinicians and anatomists.

Rosuvastatin inhibits high glucose-stimulated upregulation of VCAM-1 via the MAPK-signalling pathway in endothelial cells.

OBJECTIVE: The aim of this study is to investigate the molecular mechanisms and effect of rosuvastatin on adhesion molecule induction in human endothelial cells under high-glucose conditions (HG). METHODS AND RESULTS: The effects of rosuvastatin on vascular cell adhesion molecule (VCAM)-1 production and pERK phosphorylation were measured in HG-induced human umbilical vein endothelial cells (HUVECs) with inhibitors targeting the mitogen-activated protein kinase (MAPK) signal pathway. HG increased levels of VCAM-1. Treatment with rosuvastatin inhibited VCAM-1 expression in a concentration- and time-dependent manner. In addition, we investigated the effects of rosuvastatin on the extracellular signal-regulated kinase (ERK) 1/2 signal pathway. Rosuvastatin completely inhibited HG-induced phosphorylation of ERK. ERK/MAPK inhibitors completely prevented the VCAM-1 inhibition effect of rosuvastatin under HG condition. CONCLUSIONS: This study demonstrated that rosuvastatin suppresses HG-induced VCAM-1 production via the MAPK signalling pathway, playing a role in the suppression of atherosclerosis.

Genetic Code Expansion by Degeneracy Reprogramming of Arginyl Codons.

The genetic code in most organisms codes for 20 proteinogenic amino acids or translation stop. In order to encode more than 20 amino acids in the coding system, one of stop codons is usually reprogrammed to encode a non-proteinogenic amino acid. Although this approach works, usually only one amino acid is added to the amino acid repertoire. In this study, we incorporated non-proteinogenic amino acids into a protein by using a sense codon. As all the codons are allocated in the universal genetic code, we destroyed all the tRNA(Arg) in a cell-free protein synthesis system by using a tRNA(Arg) -specific tRNase, colicin D. Then by supplementing the system with tRNACCU , the translation system was partially restored. Through this creative destruction, reprogrammable codons were successfully created in the system to encode modified lysines along with the 20 proteinogenic amino acids.

Asymmetries in children's production of relative clauses: data from English and Korean.

We report here on a series of elicited production experiments that investigate the production of indirect object and oblique relative clauses by monolingual child learners of English and Korean. Taken together, the results from the two languages point toward a pair of robust asymmetries: children manifest a preference for subject relative clauses over indirect object relative clauses, and for direct object relative clauses over oblique relative clauses. We consider various possible explanations for these preferences, of which the most promising seems to involve the requirement that the referent of the head noun be easily construed as what the relative clause is about.

Effects of chondrocyte-derived extracellular matrix in a dry eye mouse model.

PURPOSE: The occurrence of repetitive dry eye is accompanied by inflammation. This study investigated the anti-inflammatory effects of chondrocyte-derived extracellular matrix (CDECM) on the cornea and conjunctiva in a dry eye mouse model. METHODS: Dry eyes were experimentally induced in 12- to 16-week-old NOD.B10.H2(b) mice (Control) via subcutaneous injections of scopolamine (muscarinic receptor blocker) and exposure to an air draft for 10 days (desiccation stress [DS] 10D group). Tear volume and corneal smoothness were measured at 3, 5, 7, and 10 days after the instillation of PBS (PBS group) or CDECM (CDECM group). The corneas and conjunctivas were sectioned and stained with hematoxylin and eosin (H&E) and periodic acid Schiff (PAS). The expression of inflammatory markers (i.e., tumor necrosis factor-α [TNF-α], matrix metalloproteinase-2 [MMP-2], MMP-9, intercellular adhesion molecule-1 [ICAM-1], and vascular cell adhesion molecule-1 [VCAM-1]) was detected by quantitative real-time (qRT)-PCR and western blotting. All data were statistically processed using SPSS version 18.0. RESULTS: The instillation of CDECM after the removal of the DS increased tear production by up to 3.0-fold, and corneal smoothness improved to 80% compared to the PBS group (p<0.05). In the CDECM group, the detachment of the corneal epithelial cells was reduced by 73.3% compared to the PBS group, and the conjunctival goblet cell density was significantly recovered to the control levels (p<0.05). The expression of inflammatory factors was decreased in the cornea and conjunctiva of the CDECM group compared to the PBS group. CONCLUSIONS: These observations suggest that CDECM induced effective anti-inflammatory improvements in the cornea and conjunctiva in this experimental model of dry eye.

Ramipril inhibits high glucose-stimulated up-regulation of adhesion molecules via the ERK1/2 MAPK signaling pathway in human umbilical vein endothelial cells.

Ramipril has recently been shown to have anti-atherogenic properties. However, the specific mechanisms underlying these effects remain unclear. The purpose of this study was to determine the effects of ramipril on induction of adhesion molecules in human umbilical vein endothelial cells (HUVECs) using high-glucose (HG) conditions and to investigate possible underlying molecular mechanisms. The effects of ramipril on expression of intercellular adhesion molecule (ICAM)-1, vascular cell adhesion molecule (VCAM)-1 production, and ERK phosphorylation were examined in HG-induced HUVECs with inhibitors targeting the mitogen-activated protein kinase (MAPK) signaling pathway. HG induced the expression of the adhesion molecules ICAM-1 and VCAM-1. Pretreatment with ramipril drastically inhibited ICAM-1 and VCAM-1 production in a time- and dose-dependent manner. Moreover, upon investigating the effects of ramipril on the MAPK/extracellular signal-regulated kinase (ERK) signaling pathway, we found that ramipril completely inhibited HG-induced phosphorylation of ERK1/2. ERK inhibitors completely prevented the inhibitory effect of HG. This study demonstrated that ramipril reduces HG-stimulated induction of ICAM-1 and VCAM-1 expression via MAPK signaling, which may be useful for inhibition of atherosclerosis.

Anti-neovascular effect of chondrocyte-derived extracellular matrix on corneal alkaline burns in rabbits.

PURPOSE: We investigated the effect of a chondrocyte-derived extracellular matrix (CDECM) on experimental corneal alkaline burns in rabbits. METHODS: Corneal neovascularization (NV) was induced by applying an 8-mm filter paper soaked in 1 N NaOH to the right central corneas of rabbits for 1 minute. Ten days later, the rabbits were randomly divided into three groups: the alkaline burn group, the CDECM transplantation group, and the human amniotic membrane (HAM) transplantation group. The left eyes were used as controls. CDECM and HAM were transplanted onto the corneal surface to completely cover the resected area and were subsequently sutured. On the 10th day after transplantation, the structural changes of the cornea were analyzed histologically. We examined the effects of CDECM on clinical NV features and on the expression of corneal NV markers. RESULTS: The alkaline burn produced significant NV and increased the corneal thickness. On day 10 after transplantation, the thickness, NV and opacity of the cornea were markedly decreased in the CDECM group (p < 0.001). However, the HAM transplantation group did not exhibit improvements in these clinical parameters, and there were no significant differences relative to the burn group. In addition, the use of CDECM improved the healing of the cornea following the alkaline burn by disrupting the corneal epithelial proliferation and reducing the fibrotic changes of the stroma. The hallmarks of NV were significantly induced in the subepithelium by the alkaline burn, and these levels were also suppressed by CDECM. The CDECM suppressed corneal NV by inhibiting nuclear factor-kappa B (NF-κB) activation by blocking the PKC and Akt signaling pathways. CONCLUSIONS: CDECM transplantation was markedly effective in healing alkali-burned corneas by modulating the translocation of NF-κB to the nucleus, thereby representing a promising material for the noninvasive treatment of ocular surface disease.

Evaluation of basal rate infusion in intravenous patient-controlled analgesia for post-cesarean section pain management: A randomized pilot study.

OBJECTIVE: Administering opioids via intravenous patient-controlled analgesia is a prevalent approach for managing postoperative pain. Nevertheless, due to concerns about opioid-related side effects and the potential for opioid tolerance, there is a growing emphasis on adopting opioid-sparing techniques for postoperative pain management. We aimed to investigate the effect of adding a basal rate infusion in fentanyl-based IVA following a cesarean section (CS). METHOD: Forty-eight patients, who received pain management through IVA after CS, were assigned randomly into 3 groups based on the background rate setting: Group 0 (0 mcg/hour, n = 16), Group 1 (15 mcg/hour, n = 16), and Group 2 (30 mcg/hour, n = 16). We assessed the impact of the basal infusion rate on opioid consumption and the visual analog scale (VAS) scores during the first 48 hours post-CS and also investigated opioid-induced side effects and the requirement for rescue analgesics in the ward during the first 48 hours after CS. RESULTS: In the initial 24 hours following CS, fentanyl consumption significantly increased in Group 2 compared with Group 0 and Group 1 (P = .037). At 24 hours, VAS scores both at rest and during movement, tended to decrease, as the basal rate increased; however, no significant differences were observed between the groups (P = .218 and 0.827, respectively). Between the first 24- and 48-hours post-CS, fentanyl consumption showed a marked increase in both Group 1 and Group 2 compared to Group 0 (P < .001). At 48 hours, the VAS scores at rest displayed a trend toward reduction; however, no significant differences between groups were evident (P = .165). Although the incidence of opioid-induced complications was noted, no statistically significant differences were recorded between groups during the initial 24 hours and subsequent 24 to 48 hours period (P = .556 and P = .345, respectively). CONCLUSION: The inclusion of a basal fentanyl infusion in the IVA protocol did not provide any advantages over an IVA devoid of a basal rate infusion in managing acute pain following CS.

Induction of indoleamine 2,3-dioxygenase 1 expression in neurons of the central nervous system through inhibition of histone deacetylases blocks the progression of experimental autoimmune encephalomyelitis.

BACKGROUND: A wide array of histone deacetylase (HDAC) inhibitors and aryl hydrocarbon receptor (AHR) agonists commonly arrest experimental autoimmune encephalomyelitis (EAE). However, it is not known whether HDAC inhibition is linked to the AHR signaling pathway in EAE. METHODS: We investigated how the pan-HDAC inhibitor SB939 (pracinostat) exerted immunoregulatory action in the myelin oligodendrocyte glycoprotein 35-55 (MOG35-55)-induced EAE mouse model by evaluating changes in of signal transducer and activator of transcription 3 (STAT3) acetylation and the expression of indoleamine 2,3-dioxygenase 1 (IDO1) and AHR in inflamed spinal cords during EAE evolution. We proved the involvement of IDO1 and the AHR in SB939-mediated immunosuppression using Ido1-/- and Ahr-/- mice. RESULTS: Administration with SB939 halted EAE progression, which depended upon IDO1 expression in neurons of the central nervous system (CNS). Our in vitro and in vivo studies demonstrated that SB939 sustained the interleukin-6-induced acetylation of STAT3, resulting in the stable transcriptional activation of Ido1. The therapeutic effect of SB939 also required the AHR, which is expressed mainly in CD4+ T cells and macrophages in CNS disease lesions. Finally, SB939 was shown to markedly reduce the proliferation of CD4+ T cells in inflamed neuronal tissues but not in the spleen or draining lymph nodes. CONCLUSIONS: Overall, our results suggest that IDO1 tryptophan metabolites produced by neuronal cells may act on AHR in pathogenic CD4+ T cells in a paracrine fashion in the CNS and that the specific induction of IDO1 expression in neurons at disease-afflicted sites can be considered a therapeutic approach to block the progression of multiple sclerosis without affecting systemic immunity.

Curcumin could prevent the development of chronic neuropathic pain in rats with peripheral nerve injury.

BACKGROUND: Peripheral nerve injury results in chronic neuropathic pain characterized by allodynia and/or spontaneous pain. It has been suggested that activation of mitogen-activated protein kinases such as extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) contribute to the neuropathic pain. OBJECTIVES: We investigated if curcumin could prevent the development of neuropathic pain in rats with chronic constriction injury (CCI) of the sciatic nerve. METHODS: The animals were divided into 3 groups. In the curcumin treatment group (n = 10), curcumin (50 mg/kg/d PO) was administered once daily from 1 day before CCI to 7 days after CCI. The rats in the sham group (n = 10) and CCI group (n = 10) received a control vehicle. The mechanical allodynia was assessed using von Frey at 1, 3, 5, and 7 days after nerve injury. Western blots were used to evaluate the levels of p-ERK, p-JNK, and phosphorylation of NR1 (p-NR1) subunits of N-methyl-D-aspartate in the spinal dorsal root ganglion. RESULTS: In the CCI group, mechanical allodynia was observed during 7 days after nerve injury. However, curcumin treatment reversed the mechanical allodynia 7 days after nerve ligation. There were no differences in the expression of p-ERK, p-JNK, and p-NR1 between the sham and curcumin groups. However, the expression of p-ERK, p-JNK, and p-NR1 in the CCI group were higher than the sham group and curcumin group, respectively (P < 0.05). CONCLUSIONS: Treatment with curcumin during the early stages of peripheral neuropathy can prevent the development of chronic neuropathic pain.

Measuring effective coverage of maternal and child health services in Cambodia: a retrospective analysis of Demographic and Health Surveys from 2005 to 2014.

OBJECTIVE: To investigate effective, quality-adjusted, coverage and inequality of maternal and child health (MCH) services to assess progress in improving quality of care in Cambodia. DESIGN: A retrospective secondary analysis using the three most recent (2005, 2010 and 2014) Demographic and Health Surveys. SETTING: Cambodia. PARTICIPANTS: 53 155 women aged 15-49 years old and 23 242 children under 5 years old across the three surveys. OUTCOME MEASURES: We estimated crude coverage, effective coverage and inequality in effective coverage for five MCH services over time: antenatal care (ANC), facility delivery and sick childcare for diarrhoea, pneumonia and fever. Quality was defined by the proportion of care seekers who received a set of interventions during healthcare visits. Effective coverage was estimated by combining crude coverage and quality. We used equiplots and risk ratios, to assess patterns in inequality in MCH effective coverage across wealth quintile, urban-rural and women's education levels and over time. RESULTS: In 2014, crude and effective coverage was 80.1% and 56.4%, respectively, for maternal health services (ANC and facility delivery) and 59.1% and 26.9%, respectively, for sick childcare (diarrhoea, pneumonia and fever). Between 2005 and 2014, effective coverage improved for all services, but improvements were larger for maternal healthcare than for sick child care. In 2014, poorer children were more likely to receive oral rehydration solution for diarrhoea than children from richer households. Meanwhile, women from urban areas were more likely to receive a postnatal check before getting discharged. CONCLUSIONS: Effective coverage has generally improved in Cambodia but efforts remain to improve quality for all MCH services. Our results point to substantial gaps in curative sick child care, a large share of which is provided by unregulated private providers in Cambodia. Policymakers should focus on improving effective coverage, and not only crude coverage, to achieve the health-related Sustainable Development Goals by 2030.

A Severe Cervical Spondylotic Myelopathy Patient Treated with Integrative Korean Medicine Including Acupuncture, Bee Venom Pharmacopuncture and Herbal Medicine: a case report.

Cervical spondylotic myelopathy (CSM) is common in elderly people and severe CSM patients are recommended to receive surgery. However, in some cases, surgery may fail to improve the patients' symptoms. An 80-year-old man diagnosed with CSM complained of right hemiplegia and right arm and leg pain with the presence of a Foley catheter, despite treatment with laminectomy and laminoplasty. Acupuncture, bee venom pharmacopuncture, and herbal medicine were administered for 129 days. As a result, manual muscle testing (MMT) and the Modified Barthel Index (MBI) improved, the pain in his right arm and leg decreased, and he was able to urinate by himself. This case report implies that integrative Korean medicine (IKM) can be an option for patients suffering from muscular weakness resulting from myelopathy.

Flavonoids inhibit high glucose-induced up-regulation of ICAM-1 via the p38 MAPK pathway in human vein endothelial cells.

Recently, several flavonoids have been shown to have cardioprotective, cancer preventive, or anti-inflammatory properties. However, the specific mechanisms underlying their protective effects remain unclear. We aimed to investigate the different effects of three representative flavonoids-hesperidin, naringin, and resveratrol-on intracellular adhesion molecule-1 (ICAM-1) induction in human umbilical vein endothelial cells (HUVECs) by using high-glucose (HG) concentrations and the possible underlying molecular mechanisms. In HG-induced HUVEC cultures, the effects of three different flavonoids on ICAM-1 production and p38 phosphorylation were examined in the presence or absence of inhibitors targeting the mitogen-activated protein kinase (MAPK) signal transduction pathway. HG stimulation of HUVECs increased the levels of the adhesion molecules ICAM-1 and endothelial selectin (E-selectin). Pretreatment with all the three flavonoids drastically inhibited ICAM-1 expression in a time-dependent manner, but did not alter VCAM-1 and E-selectin expressions. Moreover, we investigated the effects of flavonoids on the MAPK signal transduction pathway, because MAPK families are associated with vascular inflammation under stress. These flavonoids did not block HG-induced phosphorylation of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), but completely inhibited the HG-induced phosphorylation of p38 MAPK. SB202190, an inhibitor of p38 MAPK, also inhibited the HG-induced enrichment of ICAM-1. This study demonstrated that hesperidin, naringin, and resveratrol reduced the HG-induced ICAM-1 expression via the p38 MAPK signaling pathway, contributing to the inhibition of monocyte adhesion to endothelial cells.

Cevimeline-induced anti-inflammatory effect through upregulations of mucins in the ocular surface of a dry eye mouse model.

This study aimed to investigate the effects of various concentrations of cevimelines (CVMs) and compare them with commercial drugs in a murine model of dry eye. The experimental mouse model used male and female NOD.B10.H2b mice over 12 weeks of age. Desiccation stress was performed at 30-40% ambient humidity, and subcutaneous injection of 0.5 mg/0.2 mL scopolamine hydrobromide was performed four times a day for 10 days. The efficacy of various concentrations of CVMs (seven experimental groups) was first evaluated, and then 2% CVM was compared with commercial drugs, such as cyclosporine A (CsA), diquafosol (DQS), and rebamipide (REB) (seven experimental groups). The clinical changes, including tear production, corneal irregularity, and fluorescein staining, were measured after the instillation of various concentrations of CVMs and commercial drugs for 0, 3, 5, 7, and 10 days. Histological changes, such as corneal detachment, conjunctival goblet cell and mucin density staining, were assessed by staining the cornea or conjunctiva with hematoxylin-eosin, periodic acid-Schiff, and alcian blue. The expression of inflammatory markers and mucin factors was detected by immunohistochemistry and immunofluorescence in the lacrimal gland, cornea, and conjunctiva. Tear production was significantly increased in the 2% CVM group and was similar to that in the DQS and REB groups (P < 0.05). The corneal smoothness and fluorescein staining score were significantly improved in the 2% CVM group and were similar to those in the REB group (P < 0.05). Corneal epithelial cells were significantly decreased in the 2% CVM group, with similar observations made in the DQS and REB groups (P < 0.05). The conjunctival goblet cells and mucin density recovered in the 2% CVM group were similar to those in the CsA and REB groups (P < 0.05). The 2% CVM group showed suppressed expression of inflammatory factors in the lacrimal gland and was comparable to that seen in the CsA and REB groups. The expression of mucin factors was upregulated in the cornea and conjunctiva of the 2% CVM group and was similar to that of the CsA and REB groups. In conclusion, administration of CVM resulted in recovery or clinical and histological improvement of the murine dry eye model, and all the observed parameters were comparable to those with commercial drugs.

Stratification of Nuclear Homogeneous Patterns on HEp-2 Cells Based on Neutrophil Nuclear Staining.

Antinuclear antibody (ANA) testing is used to diagnose systemic autoimmune rheumatic disease (SARD). Nuclear homogeneous patterns on ANA-HEp-2 cells can result from anti-double-stranded DNA (dsDNA), anti-nucleosome, anti-histone, anti-Scl-70, or anti-dense fine speckles 70 (DFS70) antibodies (Abs). This study aimed to find a way to discriminate DFS70 Abs from others by way of assessing neutrophil nuclear staining on anti-neutrophil cytoplasmic antibody (ANCA) testing. Nuclear staining on ANCA-neutrophils was assessed to stratify nuclear homogeneous patterns on ANA-HEp-2 cells. Enrolled subjects included (1) young individuals with a dense fine speckled pattern on ANA testing (young non-SARD group, n=71) and patients with (2) systemic lupus erythematosus (SLE group, n=35); (3) rheumatoid arthritis possibly with histone, nucleosome Abs, and others (RA group, n=51); and (4) diffuse systemic sclerosis with Scl-70 Abs (diffuse SSc group, n=19). Negative rates (95% confidence interval) of neutrophil nuclear staining were 97.2% (90.2%-99.7%) in the young non-SARD group, 2.9% (0.1%-14.9%) in the SLE group, 3.9% (0.5%-13.5%) in the RA group, and 47.4% (24.5%-71.1%) in the diffuse SSc group. The negative rate of the young non-SARD group was significantly higher than those of the other groups (all p<0.05). In conclusion, this study suggests that the assessment of nuclear staining on ANCA-neutrophils can help to stratify nuclear homogeneous patterns on ANA-HEp-2 cells and thus to determine whether the ANA pattern is attributed to DFS70 Abs, which can be found in healthy individuals, especially in young individuals.