Plasma N-Cleaved Galectin-9 Is a Surrogate Marker for Determining the Severity of COVID-19 and Monitoring the Therapeutic Effects of Tocilizumab.

Galectin-9 (Gal-9) is known to contribute to antiviral responses in coronavirus disease 2019 (COVID-19). Increased circulating Gal-9 in COVID-19 is associated with COVID-19 severity. In a while, the linker-peptide of Gal-9 is susceptible to proteolysis that can cause the change or loss of Gal-9 activity. Here, we measured plasma levels of N-cleaved-Gal9, which is Gal9 carbohydrate-recognition domain at the N-terminus (NCRD) with attached truncated linker peptide that differs in length depending on the type of proteases, in COVID-19. We also investigated the time course of plasma N-cleaved-Gal9 levels in severe COVID-19 treated with tocilizumab (TCZ). As a result, we observed an increase in plasma N-cleaved-Gal9 levels in COVID-19 and its higher levels in COVID-19 with pneumonia compared to the mild cases (healthy: 326.1 pg/mL, mild: 698.0 pg/mL, and with pneumonia: 1570 pg/mL). N-cleaved-Gal9 levels were associated with lymphocyte counts, C-reactive protein (CRP), soluble interleukin-2 receptor (sIL-2R), D-dimer, and ferritin levels, and ratio of percutaneous oxygen saturation to fraction of inspiratory oxygen (S/F ratio) in COVID-19 with pneumonia and discriminated different severity groups with high accuracy (area under the curve (AUC): 0.9076). Both N-cleaved-Gal9 and sIL-2R levels were associated with plasma matrix metalloprotease (MMP)-9 levels in COVID-19 with pneumonia. Furthermore, a decrease in N-cleaved-Gal9 levels was associated with a decrease of sIL-2R levels during TCZ treatment. N-cleaved-Gal9 levels showed a moderate accuracy (AUC: 0.8438) for discriminating the period before TCZ from the recovery phase. These data illustrate that plasma N-cleaved-Gal9 is a potential surrogate marker for assessing COVID-19 severity and the therapeutic effects of TCZ.

Evaluation of a rapid diagnostic test for detection of dengue infection using a single-tag hybridization chromatographic-printed array strip format.

A dipstick DNA chromatography assay, a single-tag hybridization-printed array strip (STH-PAS), was evaluated for its efficacy to detect dengue virus (DENV). Reverse-transcribed DNA was amplified by PCR, and the amplified DNA was detected using the STH-PAS system. The method was evaluated using stored RNA samples previously identified to carry all 4 serotypes of dengue, chikungunya, and influenza viruses. Clinical performance was also assessed in a prospective study using plasma from 269 febrile cases from the Emergency Department of St. Luke's Medical Center, Quezon City, Philippines, and 30 afebrile normal healthy volunteers. A Taqman real-time PCR (RT-PCR) assay and a rapid Dengue NS1 test, SD Bioline, were used for comparison. The STH-PAS system was more sensitive in detecting dengue infection compared to Taqman RT-PCR. For DENV serotypes 1, 2, and 3, the detection was 1 to 2 dilutions (10-fold) higher, and for DENV serotype 4, the detection was 2-4 dilutions higher. In clinical studies, the STH-PAS system showed 100% sensitivity with 88.9% and 86.6% specificities compared to Taqman RT-PCR and SD Dengue Duo NS1 test, respectively. The STH-PAS system was found to have a superior sensitivity than the Taqman system. Further evaluation of its performance in the field may provide important data to extend its usefulness for surveillance and epidemiological research in outbreak situations.

Synthesis of a Cleaved Form of Osteopontin by THP-1 Cells and Its Alteration by Phorbol 12-Myristate 13-Acetate and BCG Infection.

The protease-cleaved osteopontin (OPN) was proposed to enhance the migration of memory T cells to granulomas in tuberculosis. Various forms of OPN were identified in human monocytic THP-1 cells stimulated by phorbol 12-myristate 13-acetate (PMA). Antibodies O-17, 10A16 and 34E3, which recognize N-terminus, the C-half, and thrombin-cleaved site of OPN, respectively, all detected distinct bands on Western blots following PMA stimulation. Bands corresponding to 18 and 30 kD were detected by antibodies 34E3 and 10A16, indicating that OPN cleavage occurred by endogenous proteases in the PMA-stimulated THP-1 cells. In immune-fluorescence (IF) assay, 34E3 positive signals were detected in intracellular space of non-infected and bacillus Calmette-Guérin (BCG)-infected cells; however, 10A16 positive signals were confirmed in extracellular area in PMA-stimulated cells followed by BCG infection. Small amounts of full-length (FL) and thrombin-cleaved (Tr) OPN were detected by ELISA in the supernatants of non-PMA-stimulated cells, and increased levels of all forms, including undefined (Ud) OPN, in PMA-stimulated cells. ELISA showed a decrease in OPN synthesis during BCG infection. To our knowledge, this is the first report of OPN cleavage in THP-1 macrophages after PMA stimulation, and of enhanced cleavage induced by BCG infection.

Comparison of antibody responses against Mycobacterium tuberculosis antigen Rv0679c in tuberculosis patients from the endemic and non-endemic regions of the Beijing genotype: a case control study.

BACKGROUND: Strains of the Beijing genotype of Mycobacterium tuberculosis (MTB) are reportedly associated with the virulence of tuberculosis (TB) infection, unfavorable outcomes of anti-TB treatment, and the global TB pandemic. Rv0679c, a hypothetical membrane protein related to host cell invasion, has a Beijing genotype-specific mutation at residue 142 (Asn142Lys). Antigenicity differences between Rv0679c-Asn142 (N-type) and Rv0679c-Lys142 (K-type) have been previously observed in mice antigen-antibody responses. However, the immune response to Rv0679c in humans remains unknown. Therefore, we aimed to investigate the anti-Rv0679c immune response in TB patients from the endemic and non-endemic regions of the Beijing MTB genotype. METHODS: We analyzed the Rv0679c-specific antibody responses in 84 subjects from the endemic region of the Beijing genotype MTB in China, including 45 pulmonary TB patients (C-PTB) and 39 healthy controls (C-HC), and 81 subjects from the Philippines (the endemic region of the non-Beijing genotype), including 51 pulmonary TB patients (P-PTB) and 30 healthy controls (P-HC). Anti-tuberculous-glycolipid (TBGL) antigen was used as the control antibody. RESULTS: TBGL IgG titers were higher in both C-PTB and P-PTB than those in their corresponding HC (C-PTB median 4.2, P-PTB median 11.2; C-PTB vs. P-PTB, p > 0.05), suggesting immune response comparability in PTB from two different countries. C-PTB showed a higher response compared to C-HC for anti-K-type IgG (53.3%) than anti-N-type IgG (6.67%); this response was not observed in P-PTB (both N-type and K-type 9.80%). CONCLUSION: Dimorphic antigen Rv0679c was found to be associated with distinct immune response patterns, indicating the role of Beijing/non-Beijing genotype of MTB in stimulating specific responses in TB patients from the endemic region of Beijing MTB. Meanwhile, reactions to Rv0679c in patients and HC from non-endemic regions of the Beijing MTB may be caused by the response to the common epitope of Rv0679c N/K-type.

Galectin-9 as a Predictive Marker for the Onset of Immune-Related Adverse Effects Associated with Anti-CCR4 MoAb Therapy in Patients with Adult T Cell Leukemia.

Adult T-cell leukemia/lymphoma (ATL/ATLL) is one of the most malignant lymphomas with poor prognosis. ATL/ATLL cells express CC chemokine receptor 4, and mogamulizumab (anti-CCR4 monoclonal antibody) exhibits strong cytotoxicity for ATL/ATLL cells. We analyzed plasma samples of 6 patients with ATL/ATLL treated with chemotherapy followed by mogamulizumab therapy (mogatherapy) for changes in the levels of biomarkers in relation to immune-related adverse effects. As treatment is often associated with skin eruptions, we investigated the profiles of inflammatory cytokines, including galectin-9 (Gal-9), which becomes increased in various infectious diseases and allergic patients. Gal-9, soluble interleukin (IL)-2 receptor, tumor necrosis factor-α, and IL-10 levels were increased before chemotherapy, and Gal-9 levels were associated with the sIL-2 receptor, which reflects tumor burden. Inflammatory levels decreased after chemotherapy. After mogatherapy, 5 of 6 patients attained complete remission (CR), whereas 1 patient showed no response (NR) and died. Among 5 patients with CR, the biomarkers remained low during mogatherapy, except for a 3-5-fold increment in Gal-9 (associated with skin eruptions). A skin biopsy showed infiltration by inflammatory cells and Gal-9 synthesis in areas with CD8 cell infiltration. In the patient with NR, increased levels of Gal-9 and the aforementioned biomarkers were noted 3 days after mogatherapy, followed by opportunistic infections resembling immune reconstitution inflammatory syndrome. Therefore, an increased Gal-9 plasma level in ATL/ATLL indicates tumor burden and reflects immune activation by mogatherapy. These findings may indicate that an increase in the Gal-9 level, a novel immune checkpoint molecule, can reflect immune-related adverse effects of various biotherapies.

Secretion of Galectin-9 as a DAMP during Dengue Virus Infection in THP-1 Cells.

Damage-associated molecular patterns (DAMPs) are endogenous cellular molecules released to the extracellular environment in response to stress conditions such as virus infection. Galectins are ß-galactoside-binding proteins that are widely expressed in cells and tissues of the immune system, are localized in the cell cytoplasm, and have roles in inflammatory responses and immune responses against infection. Elevated levels of galectin-9 (Gal-9) in natural human infections have been documented in numerous reports. To investigate the effect of dengue virus (DENV) infection on expression of endogenous Gal-9, monocytic THP-1 cells were infected with varying doses of DENV-3 (multiplicity of infection (MOI) 0.01, 0.03 and 0.1) and incubated at varying time points (Day 1, Day 2, Day 3). Results showed augmentation of Gal-9 levels in the supernatant, reduction of Gal-9 levels in the cells and decreased expression of LGALS9 mRNA, while DENV-3 mRNA copies for all three doses remained stable through time. Dengue virus induced the secretion of Gal-9 as a danger response; in turn, Gal-9 and other inflammatory factors, and stimulated effector responses may have limited further viral replication. The results in this pilot experiment add to the evidence of Gal-9 as a potential DAMP.

Secretion of IFN-γ Associated with Galectin-9 Production by Pleural Fluid Cells from a Patient with Extrapulmonary Tuberculosis.

In this study, we investigated the role of a matricellular protein galectin-9 (Gal-9) in pleural effusion related to tuberculosis (TB). Plasma and pleural fluid of a patient with extrapulmonary TB were analyzed for cytokine content by ELISA and Luminex. Peripheral blood mononuclear cells (PBMCs) and pleural fluid cells (PFCs) were examined for interferon-γ (IFN-γ) secretion by the enzyme-linked immunospot (ELISPOT) assay or IFN-γ ELISA, for apoptosis and necrosis by Cell Death Detection ELISA, and also underwent cell sorting. The results indicate that compared to plasma, pleural fluid had increased levels of IFN-γ (1.6 vs. 55.5 pg/mL), IL-10, IL-12p40, vascular endothelial growth factor (VEGF), and Gal-9 (3.0 vs. 936.0 pg/mL), respectively. PFCs culture supernatant exhibited higher concentration of Gal-9 compared to PBMCs in culture, consistent with enriched Gal-9 staining in the granuloma that is in closer vicinity to PFCs compared to PBMCs. PFCS displayed higher IFN-γ secretion after stimulation with TB antigens ESAT-6/CFP-10. Furthermore, in PFCs, Gal-9 alone could stimulate IFN-γ synthesis in culture or ELISPOT, which was inhibited by a Gal-9 antagonist lactose, and which may promote apoptosis and necrosis. These findings suggest that Gal-9 could modulate immune responses and participate in immunopathology of pleural effusion during TB.

Plasma levels of Galectin-9 reflect disease severity in malaria infection.

BACKGROUND: Galectin-9 (Gal-9) is a ß-galactoside-binding lectin that interacts with sugar moieties on glycoproteins and glycolipids of cells and pathogens. Gal-9 is known as an immune modulator that induces cell death via interaction with T cell immunoglobulin and mucin domain-3 (Tim3), a co-inhibitory receptor, and it inhibits production of several pro-inflammatory cytokines (TNF, IL-6 and IL-1α) and enhances production of IL-10. To understand the immune pathology of malaria, the Gal-9 in plasma was measured. METHODS: Plasma samples and clinical parameters were obtained from 50 acute malaria cases (nine severe and 41 uncomplicated cases) from Thailand at three time points: day 0, day 7 and day 28. Gal-9 levels were determined by ELISA. A total of 38 species of cytokines and chemokines were measured using a BioPlex assay. RESULTS: Gal-9 levels were higher at day 0 compared to day 7 and day 28 (P < 0.0001). Gal-9 levels were also higher in severe malaria (SM) cases compared to uncomplicated (UM) cases at day 0 and day 7 (923 vs 617 pg/mL; P = 0.03, and 659 vs 348 pg/mL; P = 0.02 respectively). Median Gal-9 levels were higher in patients with blood urea nitrogen to creatinine ratio (BUN/creatinine) ≥20 (mg/dL) than in patients with BUN/creatinine <20 (mg/dL) at day 0 (817.3 vs 576.2 pg/mL, P = 0.007). Gal-9 was inversely significantly correlated with chloride levels in both SM and UM cases (r s = -0.73 and r s = -0.46, respectively). In both UM and SM cases, Gal-9 was significantly associated with pro- and anti-inflammatory cytokines and chemokines such as TNF, IL-6, IFN-α2, IFN-γ, IL-1Ra and IL-10. These correlations were observed at day 0 but disappeared at day 28. CONCLUSIONS: Gal-9 is released during acute malaria, and reflects its severity. This elevation of Gal-9 in acute malaria infection raises the possibility of its role in termination of the immune response by binding to Tim-3, a receptor of Gal-9.

Prevention of Tetanus Outbreak Following Natural Disaster in Indonesia: Lessons Learned from Previous Disasters.

In Indonesia, the Aceh earthquake and tsunami in 2004 killed 127,000 people and caused half a million injuries, while the Yogyakarta earthquake in 2006 caused 5,700 deaths and 37,000 injuries. Because disaster-affected areas are vulnerable to epidemic-prone diseases and tetanus is one such disease that is preventable, we systematically reviewed the literature related to tetanus outbreaks following previous two natural disasters in Indonesia. Based on our findings, recommendations for proper vaccination and education can be made for future countermeasures. Using specified keywords related to tetanus and disasters, relevant documents were screened from PubMed, the WHO website, and books. Reports offering limited data and those released before 2004 were excluded. In all, 16 publications were reviewed systematically. Results show that 106 cases of tetanus occurred in Aceh, with a case fatality ratio (CFR) of 18.9%; 71 cases occurred in Yogyakarta, with CFR of 36.6%. For both outbreaks, most patients had been wounded during scavenging or evacuation after the disaster occurred. Poor access to health care because of limited transportation or hospital facilities, and low vaccination coverage and lack of awareness of tetanus risk contributed to delayed treatment and case severity. Tetanus outbreaks after disasters are preventable by increasing vaccination coverage, improving wound care treatment, and establishing a regular surveillance system, in addition to good practices of disaster management and supportive care following national guidelines. Furthermore, health education for communities should be provided to raise awareness of tetanus risk reduction.

Development of Point-of-Care Testing for Disaster-Related Infectious Diseases.

After disaster, the victims lose their safe lives and are even exposed to nature where they could suffer from animal bites and vectors followed by suffering from zoonosis or vector-born diseases. Because of the urgent need for rapid and cheap diagnosis for infectious diseases after disaster, anonymous questionnaire clarified that leptospirosis, dengue, diarrhea, and cholera were recognized as common disaster-related infections in the Philippines, while diarrhea and pneumonia were more common in Indonesia. It should also be noted that infectious disease itself such as tuberculosis associated with acquired immune deficiency syndrome in South Africa is a disaster. Thus, the possible occurrence of similar situation in Asia should be prevented. We have conducted an international collaborative research in the Philippines and Indonesia on dengue virus, leptospira and mycobacterium tuberculosis (MTB) infectious diseases. Development of point-of-care testing for molecular diagnosis and disease severity was the principal purpose of the research. Loop-mediated isothermal amplification assay, which does not require a source of electricity, was developed for leptospirosis, dengue and MTB and has been proved to be useful where resource is limited. The plasma levels of matricellular proteins, including galectin-9 and osteopontin, were found to reflect the disease severities in dengue virus and MTB infection, probably because matricellular proteins are one of the most functional extracellular proteins that are associated with inflammatory edema. The study on disaster-related infectious disease facilitates the international cooperation for development of point-of-care testing for tropical infectious diseases.

Immunological Roles of Elevated Plasma Levels of Matricellular Proteins in Japanese Patients with Pulmonary Tuberculosis.

Elevated matricellular proteins (MCPs), including osteopontin (OPN) and galectin-9 (Gal-9), were observed in the plasma of patients with Manila-type tuberculosis (TB) previously. Here, we quantified plasma OPN, Gal-9, and soluble CD44 (sCD44) by enzyme-linked immunosorbent assay (ELISA), and another 29 cytokines by Luminex assay in 36 patients with pulmonary TB, six subjects with latent tuberculosis (LTBI), and 19 healthy controls (HCs) from Japan for a better understanding of the roles of MCPs in TB. All TB subjects showed positive results of enzyme-linked immunospot assays (ELISPOTs). Spoligotyping showed that 20 out of 36 Mycobacterium tuberculosis (MTB) strains belong to the Beijing type. The levels of OPN, Gal-9, and sCD44 were higher in TB (positivity of 61.1%, 66.7%, and 63.9%, respectively) than in the HCs. Positive correlations between OPN and Gal-9, between OPN and sCD44, and negative correlation between OPN and ESAT-6-ELISPOT response, between chest X-ray severity score of cavitary TB and ESAT-6-ELISPOT response were observed. Instead of OPN, Gal-9, and sCD44, cytokines G-CSF, GM-CSF, IFN-α, IFN-γ, IL-12p70, and IL-1RA levels were higher in Beijing MTB-infected patients. These findings suggest immunoregulatory, rather than inflammatory, effect of MCPs and can advance the understanding of the roles of MCPs in the context of TB pathology.

Urine Levels of Defensin α1 Reflect Kidney Injury in Leptospirosis Patients.

Leptospirosis is a zoonotic disease whose severe forms are often accompanied by kidney dysfunction. In the present study, urinary markers were studied for potential prediction of disease severity. Urine samples from 135 patients with or without leptospirosis at San Lazaro Hospital, the Philippines, were analyzed. Urine levels of defensin α1 (uDA1) were compared with those of neutrophil gelatinase-associated lipocalin (uNGAL) and N-acetyl-ß-d-glucosidase (uNAG). Serum creatinine (Cr) was used as a marker of kidney injury. The levels of uDA1/Cr, uNGAL/Cr, and uNAG/Cr were positive in 46%, 90%, and 80% of leptospirosis patients, and 69%, 70%, and 70% of non-leptospirosis patients, respectively. In leptospirosis patients, the correlation of uDA1/Cr, uNGAL/Cr and uNAG/Cr levels with serum Cr were r = 0.3 (p < 0.01), r = 0.29 (p < 0.01), and r = 0.02 (p = 0.81), respectively. uDA1/Cr levels were correlated with uNGAL/Cr levels (r = 0.49, p < 0.01) and uNAG/Cr levels (r = 0.47, p < 0.0001) in leptospirosis patients. These findings suggest that uDA1, uNGAL, and uNAG were elevated in leptospirosis patients and reflected various types of kidney damage. uDA1 and uNGAL can be used to track kidney injury in leptospirosis patients because of their correlation with the serum Cr level.

Osteopontin-integrin interaction as a novel molecular target for antibody-mediated immunotherapy in adult T-cell leukemia.

BACKGROUND: Adult T-cell leukemia (ATL) is a CD4(+) T-cell neoplasm with a poor prognosis. A previous study has shown that there is a strong correlation between the secreted matricellular protein osteopontin (OPN) level and disease severity in ATL patients. Here, we investigated the role of OPN in ATL pathogenesis and the possible application of anti-OPN monoclonal antibody (mAb) for ATL immunotherapy in NOD/Shi-scid,IL-2Rg (null) (NOG) mice. RESULTS: Subcutaneous inoculation of ATL cell lines into NOG mice increased the plasma level of OPN, which significantly correlated with metastasis of the inoculated cells and survival time. Administration of an SVVYGLR motif-recognizing anti-OPN mAb resulted in inhibition not only of tumor growth but also of tumor invasion and metastasis. The number of fibroblast activating protein-positive fibroblasts was also reduced by this mAb. We then co-inoculated mouse embryonic fibroblasts (MEFs) isolated from wild-type (WT) or OPN knockout mice together with ATL-derived TL-OmI cells into the NOG mice. The mice co-inoculated with WT MEFs displayed a significant decrease in survival relative to those injected with TL-OmI cells alone and the absence of OPN in MEFs markedly improved the survival rate of TL-OmI-inoculated mice. In addition, tumor volume and metastasis were also reduced in the absence of OPN. CONCLUSION: We showed that the xenograft NOG mice model can be a useful system for assessment of the physiological role of OPN in ATL pathogenesis. Using this xenograft model, we found that fibroblast-derived OPN was involved in tumor growth and metastasis, and that this tumor growth and metastasis was significantly suppressed by administration of the anti-OPN mAbs. Our findings will lead to a novel mAb-mediated immunotherapeutic strategy targeting against the interaction of OPN with integrins on the tumor of ATL patients.

Evaluation of matricellular proteins in systemic and local immune response to Mycobacterium tuberculosis infection.

Matricellular proteins such as osteopontin (OPN), galectin-9 (Gal-9), and tenascin-C (TN-C) are expressed not only under normal physiological conditions, but also during infection, inflammation and tumorigenesis. Plasma concentrations of matricellular proteins were studied to determine their diagnostic value as potential markers of tuberculosis (TB) activity. It was found that concentrations of OPN and TN-C were higher in patients with active TB than in healthy controls and individuals with latent infection. Moreover, LTBI patients had higher concentrations of OPN than did healthy controls. Gal-9 concentrations did not differ significantly between groups. Concentrations of matricellular proteins were higher in pleural fluid than in the plasma of patients with TB. Expression of matricellular proteins was also investigated in TB granulomas and other granulomatous diseases. Positive OPN and Gal-9 staining was observed in TB and sarcoidosis granulomas, but not in Crohn disease granulomas. The fibrotic ring around granulomas stained positive for TN-C in TB and sarcoidosis, but not in Crohn disease. Of the three matricellular proteins studied, OPN and TN-C may serve as reliable plasma markers for monitoring TB activity, whereas Gal-9 seems to be expressed more at the site of infection than in the systemic circulation.

Leptospirosis in the Tohoku region: re-emerging infectious disease.

Leptospirosis is a zoonotic and disaster-related infectious disease. It is mainly endemic in subtropical or tropical countries and has not been reported since 2009 in the Tohoku region (northern Japan), including the Yamagata and Miyagi Prefectures. However, we experienced four patients with leptospirosis in the Tohoku region from 2012 to 2014; three patients (#1-3) live in the agricultural areas of the Yamagata Prefecture and one patient (#4) was a visitor to the Miyagi Prefecture. Patient 1 (81-year-old female) is a villager, with a rat bite, while Patient 2 (77-year-old male) and Patient 3 (84-year-old female) are farmers and were infected probably during agriculture work. Patient 4 (40-year-old male US citizen) was infected while traveling in Thailand. They had chief complaint of fever, headache, and myalgia and showed manifestations of hyperbilirubinemia (mean, 4.35 mg/dL), thrombocytopenia and acute kidney injury (AKI). All patients were diagnosed by polymerase chain reaction using blood and/or urine samples and a microscopic agglutination test for the anti-Leptospira antibody. All the patients were treated with infused antibiotics, including minocycline. The patients underwent hemodialysis due to severe AKI (mean serum creatinine, 4.44 mg/dL), except for Patient 2 with the normal serum creatinine level (1.12 mg/dL). All the patients recovered and were discharged. The presence of the three patients in the Yamagata Prefecture implies that leptospirosis does re-emerge in the Tohoku region. Therefore, careful survey of the pathogen is necessary for febrile patients with AKI who engage in agriculture or have a recent history of travelling in subtropical or tropical countries.

Elevated OPN, IP-10, and neutrophilia in loop-mediated isothermal amplification confirmed tuberculosis patients.

Tuberculosis (TB) is the second most common cause of death from infectious diseases and results in high socioeconomic losses to many countries. Proper diagnosis is the first step in TB eradication. To develop a rapid, simple, and accurate diagnostic TB test and to characterize the prevalence of Mycobacterium tuberculosis (MTB) genotypes and immune profiles of TB patients, a total of 37 TB patients and 30 healthy control (HC) from Metro Manila were enrolled. Loop-mediated isothermal amplification (LAMP) reliably detected MTB infection. Manila genotype was identified by spoligotyping method in all TB patients. Osteopontin (OPN), interferon-γ-induced protein 10 kDa (IP-10), and neutrophil counts were found to reflect the acute stage of MTB infection. The sensitivity and specificity were 94.6% and 93.3%, respectively, for both OPN and IP-10, and they were 83.8% and 78.6%, respectively, for neutrophils. The combination of OPN, IP-10, neutrophil count, IL-6, IL-8, TNF-α, MCP-1, platelets, galectin-9, and leukocyte count correctly identifies all the HC and 96.3% of TB patients. LAMP method may serve as a rapid, supportive method in addition to time-consuming culture methods. OPN, IP-10, and neutrophil counts are useful in detecting MTB infection and may have utility in monitoring the course of the disease.

The elevation of plasma galectin-9 levels in patients with psoriasis and its associations with inflammatory and immune checkpoint molecules in skin tissues.

Psoriasis is a chronic, immune-mediated disorder that mainly affects the skin, with an estimated global prevalence of 2-3%. Galectin-9 (Gal-9) is a ß-galactoside-binding lectin capable of promoting or suppressing the progression of infectious and immune-mediated diseases. Here, we determined if the expression of Gal-9 is observed in psoriasis. Gal-9 levels were measured in plasma of psoriasis (n = 62) and healthy control (HC) (n = 31) using an enzyme-linked immunosorbent assay. In addition, skin samples from seven patients were screened for RNA transcriptomes and the expression of Gal-9 was compared with inflammatory, immune checkpoint molecules (ICMs) and Foxp3. The plasma Gal-9 levels in patients with psoriasis were significantly higher (841 pg/mL) than in HCs (617 pg/mL) (P < 0.0001) and were associated with white blood cell numbers, eosinophils (%) and alanine transaminase. The levels of inflammatory molecules IL-36B, IL-17RA, IL-6R, IL-10, IRF8, TGFb1, and IL-37, and those of ICMs of Tim-3, CTLA-4, CD86, CD80, PD-1LG2, CLEC4G, and Foxp3 were significantly correlated with Gal-9 (LGALS9) in skin. However, HMGB1, CD44, CEACAM1 and PDL1-known to be associated with a variety of Gal-9 biological functions were not correlated with LGALS9. Thus, it is likely that Gal-9 expression affects the disease state of PS.

Quinolone-induced upregulation of osteopontin gene promoter activity in human lung epithelial cell line A549.

Quinolones, in addition to their antibacterial activities, act as immunomodulators. Osteopontin (OPN), a member of the extracellular matrix proteins, was found to play a role in the immune and inflammatory response. We found that quinolones significantly enhanced OPN secretion, namely, garenoxacin (220%), moxifloxacin (62%), gatifloxacin (82%), sparfloxacin, (79%), and sitafloxacin (60%). Enhancement of OPN secretion was shown to be due to the effect of quinolones on the OPN gene promoter activity. We also examined the role of quinolones on apoptosis and found that sparfloxacin decreased the late apoptosis of A549 cells, but garenoxacin did not show the antiapoptotic effect. The antiapoptotic effects of quinolones do not appear to be associated with OPN elevation.

Frequent detection of anti-tubercular-glycolipid-IgG and -IgA antibodies in healthcare workers with latent tuberculosis infection in the Philippines.

Anti-tubercular-glycolipid-IgG (TBGL-IgG) and -IgA (TBGL-IgA) antibodies, and the QuantiFERON-TB Gold test (QFT) were compared in healthcare workers (HCWs, n = 31) and asymptomatic human immunodeficiency virus-carriers (HIV-AC, n = 56) in Manila. In HCWs, 48%, 51%, and 19% were positive in QFT, TBGL-IgG, and -IgA, respectively. The TBGL-IgG positivity was significantly higher (P = 0.02) in QFT-positive than QFT-negative HCWs. Both TBGL-IgG- and -IgA-positive cases were only found in QFT-positive HCWs (27%). The plasma IFN-γ levels positively correlated with TBGL-IgA titers (r = 0.74, P = 0.005), but not TBGL-IgG titers in this group, indicating that mucosal immunity is involved in LTBI in immunocompetent individuals. The QFT positivity in HIV-AC was 31% in those with CD4+ cell counts >350/µL and 12.5% in low CD4 group (<350/µL). 59 % and 29% were positive for TBGL-IgG and -IgA, respectively, in HIV-AC, but no association was found between QFT and TBGL assays. TBGL-IgG-positive rates in QFT-positive and QFT-negative HIV-AC were 61% and 58%, and those of TBGL-IgA were 23% and 30%, respectively. The titers of TBGL-IgA were associated with serum IgA (P = 0.02) in HIV-AC. Elevations of TBGL-IgG and -IgA were related to latent tuberculosis infection in HCWs, but careful interpretation is necessary in HIV-AC.

Rapid decrease of plasma galectin-9 levels in patients with acute HIV infection after therapy.

Acute HIV-1 infection is often diagnosed as infectious mononucleosis and the symptoms resolve spontaneously after varying periods of time. After the infection of HIV-1 through the mucosa, the characteristic clinical symptoms and laboratory markers of acute HIV-1 infection appear in each patient through a complicated virus-host interaction. To understand the host responses, we measured two unique proinflammatory cytokines, galectin-9 (Gal-9) and osteopontin (OPN). A ß-galactoside-binding mammalian lectin, Gal-9, reduces pro-inflammatory type-1 helper T (Th1) cells and Th17 cells and increases anti-inflammatory regulatory T cells. The plasma level of Gal-9 is known to be associated with HIV-1 viral load in chronic HIV-1 infection. On the contrary, osteopontin induces Th1/Th17 cells and promotes tissue inflammation. OPN is synthesized by variety of cells in the body, and dendritic cells are known to synthesize OPN in HIV-1 infected individuals. It was hypothesized that Gal-9 and/or OPN could be not only immune-modulators but also novel biomarkers of acute HIV-1 infection. We experienced 3 patients with acute HIV-1 and measured the levels of Gal-9 and OPN periodically before and after antiretroviral treatment. The results showed that the plasma levels of Gal-9 were extremely elevated [more than 2,300 pg/ml (normal range < 46 pg/ml)] in all three acute HIV-1 infected individuals and decreased rapidly after treatment. The changes in the OPN levels were less marked. In conclusion, the plasma levels of Gal-9 may be predictive of a severe inflammation status during the acute phase of HIV-1 infection and could be a potential biomarker during acute infection.