High incidence of serious infections requiring hospitalisation in human T-cell leukaemia virus type 1-positive rheumatoid arthritis: A case-controlled observational study.

OBJECTIVES: We aimed to assess the clinical features of human T-cell leukaemia virus type 1 (HTLV-1)-positive rheumatoid arthritis (RA) patients. Furthermore, we investigated the impact of HTLV-1 infection on incidences of serious infections requiring hospitalisation (SIH) and malignancies. METHODS: A total of 150 sex- and age-matched HTLV-1-negative and 50 HTLV-1-positive RA patients were enrolled from the HTLV-1 RA Miyazaki Cohort Study. Clinical and laboratory data were collected from this cohort database. The incidence rate (IR) for SIH and malignancies from 2015 to 2020 was analysed. RESULTS: The median age and female ratio in the study population were 70 years old and 80%, respectively. Although no differences were found in inflammatory marker values between the two groups, the patient global assessment and Health Assessment Questionnaire scores were higher in HTLV-1-positive RA patients. In HTLV-1-negative RA patients, the IR for SIH was 6.37/100 person-years (PY) and 1.32/100 PY for malignancies. In HTLV-1-positive RA patients, SIH occurred in 11.1/100 PY and malignancies in 2.46/100 PY. The crude IR ratio comparing SIH between two groups was 1.74 (95% confidence interval, 1.04-2.84), which was a significant increase. CONCLUSIONS: HTLV-1-positive RA patients may worsen RA symptoms. HTLV-1 may be a risk factor for SIH.

Clinical significance of soluble CADM1 as a novel marker for adult T-cell leukemia/lymphoma.

Adult T-cell leukemia/leukemia (ATLL) is an aggressive peripheral T-cell malignancy, caused by infection with the human T-cell leukemia virus type 1 (HTLV-1). We have recently shown that cell adhesion molecule 1 (CADM1), a member of the immunoglobulin superfamily, is specifically and consistently overexpressed in ATLL cells, and functions as a novel cell surface marker. In this study, we first show that a soluble form of CADM1 (sCADM1) is secreted from ATLL cells by mainly alternative splicing. After developing the Alpha linked immunosorbent assay (AlphaLISA) for sCADM1, we showed that plasma sCADM1 concentrations gradually increased during disease progression from indolent to aggressive ATLL. Although other known biomarkers of tumor burden such as soluble interleukin-2 receptor α (sIL-2Rα) also increased with sCADM1 during ATLL progression, multivariate statistical analysis of biomarkers revealed that only plasma sCADM1 was selected as a specific biomarker for aggressive ATLL, suggesting that plasma sCADM1 may be a potential risk factor for aggressive ATLL. In addition, plasma sCADM1 is a useful marker for monitoring response to chemotherapy as well as for predicting relapse of ATLL. Furthermore, the change in sCADM1 concentration between indolent and aggressive type ATLL was more prominent than the change in the percentage of CD4+CADM1+ ATLL cells. As plasma sCADM1 values fell within normal ranges in HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) patients with higher levels of serum sIL-2Rα, a measurement of sCADM1 may become a useful tool to discriminate between ATLL and other inflammatory diseases, including HAM/TSP.

Effectiveness and safety of non-tumor necrosis factor inhibitor therapy for anti-human T-cell leukemia virus type 1 antibody-positive rheumatoid arthritis.

OBJECTIVES: Our previous study showed that the effectiveness of tumor necrosis factor (TNF) inhibitors was attenuated in anti-human T-cell leukemia virus type 1 (HTLV-1) antibody-positive patients with rheumatoid arthritis (RA). We aimed to evaluate the effectiveness and safety of non-TNF inhibitors in anti-HTLV-1 antibody-positive patients with RA. METHODS: We reviewed patients with RA who received abatacept or tocilizumab as the first biologic agent. We used the data of patients treated with TNF inhibitors from our previous study to compare the effectiveness between the anti-HTLV-1 antibody-positive patients treated with TNF inhibitors and non-TNF inhibitors using the inverse probability of treatment weights (IPTW) method. RESULTS: A total of 359 patients were divided into anti-HTLV-1 antibody-negative and -positive patients of 332 and 27, respectively. No statistically significant difference was observed in the change in the clinical disease activity index between the anti-HTLV-1 antibody-positive and -negative patients. The results using the IPTW method showed a significant association between the non-TNF inhibitors treatment and a better response. None of the patients developed adult T-cell leukemia/lymphoma or HTLV-1-associated myelopathy/tropical spastic paraparesis during the 24 weeks. CONCLUSION: Our results indicate that non-TNF inhibitors treatment is safety, and the effectiveness is not attenuated also in anti-HTLV-1 antibody-positive patients.

Establishment of a novel diagnostic test algorithm for human T-cell leukemia virus type 1 infection with line immunoassay replacement of western blotting: a collaborative study for performance evaluation of diagnostic assays in Japan.

BACKGROUND: The reliable diagnosis of human T-cell leukemia virus type 1 (HTLV-1) infection is important, particularly as it can be vertically transmitted by breast feeding mothers to their infants. However, current diagnosis in Japan requires a confirmatory western blot (WB) test after screening/primary testing for HTLV-1 antibodies, but this test often gives indeterminate results. Thus, this collaborative study evaluated the reliability of diagnostic assays for HTLV-1 infection, including a WB-based one, along with line immunoassay (LIA) as an alternative to WB for confirmatory testing. RESULTS: Using peripheral blood samples from blood donors and pregnant women previously serologically screened and subjected to WB analysis, we analyzed the performances of 10 HTLV-1 antibody assay kits commercially available in Japan. No marked differences in the performances of eight of the screening kits were apparent. However, LIA determined most of the WB-indeterminate samples to be conclusively positive or negative (an 88.0% detection rate). When we also compared the sensitivity to HTLV-1 envelope gp21 with that of other antigens by LIA, the sensitivity to gp21 was the strongest. When we also compared the sensitivity to envelope gp46 by LIA with that of WB, LIA showed stronger sensitivity to gp46 than WB did. These findings indicate that LIA is an alternative confirmatory test to WB analysis without gp21. Therefore, we established a novel diagnostic test algorithm for HTLV-1 infection in Japan, including both the performance of a confirmatory test where LIA replaced WB on primary test-reactive samples and an additional decision based on a standardized nucleic acid detection step (polymerase chain reaction, PCR) on the confirmatory test-indeterminate samples. The final assessment of the clinical usefulness of this algorithm involved performing WB analysis, LIA, and/or PCR in parallel for confirmatory testing of known reactive samples serologically screened at clinical laboratories. Consequently, LIA followed by PCR (LIA/PCR), but neither WB/PCR nor PCR/LIA, was found to be the most reliable diagnostic algorithm. CONCLUSIONS: Because the above results show that our novel algorithm is clinically useful, we propose that it is recommended for solving the aforementioned WB-associated reliability issues and for providing a more rapid and precise diagnosis of HTLV-1 infection.

Neutrophil-derived lactoferrin induces the inflammatory responses of rheumatoid arthritis synovial fibroblasts via Toll-like receptor 4.

OBJECTIVES: Damage-associated molecular patterns (DAMPs) are proposed to drive aberrant stimulation of Toll-like receptors (TLRs) in rheumatoid arthritis (RA) inflamed joints. In the current study we investigated the role of the neutrophil-derived lactoferrin (LTF), as an endogenous ligand for TLR4 in the inflammatory response of RA synovial fibroblasts (RASFs). METHODS: RASFs were stimulated with LTF, and the expressions of inflammatory cytokines in RASFs were measured. To clarify the TLR4 signalling pathway associated with LTF stimulation, a small molecular inhibitor of TLR4 (TAK242) and NF-κB inhibitor were used. The role of nuclear factor of activated T cells 5 (NFAT5) was identified using small interfering RNA. To reveal the interaction between NF-κB and NFAT5, cerulenin, which disrupts their interaction, was used. RESULTS: Stimulation of RASFs with LTF significantly increased the expressions of inflammatory cytokines and chemokines, such as IL-6, CCL20 and IL-8, in RASFs. LTF enhanced the mRNA expressions of these cytokines in RASFs stimulated by TNF-α. TAK242 almost completely inhibited the expressions of inflammatory cytokines and chemokines in RASFs stimulated by LTF. The NF-κB inhibitor partially repressed the expressions of IL-6 and IL-8 mRNAs induced by LTF, but not CCL20 mRNA expression. On the other hand, NFAT5 silencing decreased the expressions of CCL20 and IL-8 mRNAs induced by LTF, but not IL-6 mRNA expression. Cerulenin repressed the expressions of IL-6, CCL20 and IL-8 in RASFs stimulated by LTF. CONCLUSIONS: Neutrophil-derived LTF may play a role as an endogenous ligand for TLR4 expressed on RASFs. NFAT5-NF-κB enhanceosome might regulate the expressions of LTF-TLR4-responsive genes in RASFs.

The time-sequential changes of risk factors for adult T-cell leukemia development in human T-cell leukemia virus-positive patients with rheumatoid arthritis: a retrospective cohort study.

Objective: This study aimed to investigate the time-sequential changes of risk factors for adult T-cell leukemia (ATL) development in human T-cell leukemia virus type 1 (HTLV-1)-positive rheumatoid arthritis (RA) patients. Methods: HTLV-1 infection was screened using particle agglutination assay and confirmed via western blotting in 365 RA patients. Twenty-three HTLV-1-positive RA patients were included in the study cohort. Blood samples were obtained from these patients at each observation time point. The values of HTLV-1 proviral load (PVL) and serum soluble IL-2 receptor (sIL2-R), which are risk factors for ATL development, were measured using real-time PCR and enzyme immunoassay, respectively. Results: The study cohort comprised 79 person-years. The median HTLV-1 PVL and sIL2-R values of the HTLV-1-positive RA patients were 0.44 copies per 100 white blood cells (WBCs) and 406 U/mL, respectively. Three HTLV-1-positive RA patients showed a high PVL value. No remarkable changes were observed in the PVL and sIL2-R values during the observation period. However, one elderly HTLV-1-positive RA patient who had a high PVL value developed ATL during treatment with methotrexate and infliximab. Conclusion: A thorough clinical assessment of the risk factors for ATL development may be necessary in daily clinical practice for RA patients in HTLV-1-endemic areas in Japan.

Development of reference material with assigned value for human T-cell leukemia virus type 1 quantitative PCR in Japan.

Quantitative PCR (qPCR) of human T-cell leukemia virus type 1 (HTLV-1) provirus is used for HTLV-1 testing and for assessment of risk of HTLV-1-related diseases. In this study, a reference material was developed for standardizing HTLV-1 qPCR. Freeze-dried TL-Om1 cells diluted with Jurkat cells were prepared and an assigned value for proviral load (PVL) of 2.71 copies/100 cells was determined by digital PCR. Nine Japanese laboratories using their own methods evaluated the PVLs of this reference material as 1.08-3.49 copies/100 cells. The maximum difference between laboratories was 3.2-fold. Correcting measured PVLs by using a formula incorporating the assigned value of this reference material should minimize such discrepancies.

Tumor necrosis factor alpha inhibitors have no effect on a human T-lymphotropic virus type-I (HTLV-I)-infected cell line from patients with HTLV-I-associated myelopathy.

BACKGROUND: While tumor necrosis factor alpha (TNF-α) inhibitors (TNFi) and other biologics are very effective against autoimmune diseases, they can also cause infectious diseases. Therefore, it is important to clarify whether the TNFi sometimes used to treat patients with rheumatoid arthritis (RA) complicated with human T-lymphotropic virus type-I (HTLV-I) infection have the unintended side effect of promoting HTLV-I proliferation. METHODS: We used the HTLV-I-infected cell line HCT-5, derived from spinal fluid cells of a patient with HTLV-I associated myelopathy, to evaluate the production of cytokines and chemokines, TNF-α receptor (TNFR), the expression of HTLV-I associated genes, the HTLV-I proviral load (PVL), the expression of HTLV-I structural protein, and apoptosis. We used Jurkat cells as a control. RESULTS: Supernatants of HCT-5 showed time-dependent elevations of IL-6, RANTES and ICAM-1. HCT-5 supernatants treated with infliximab, adalimumab, etanercept (ETN), golimumab and certolizumab pegol showed no significant differences in the levels of these molecules compared to the control. Neither TNFR1 nor TNFR2 expression was altered by any TNFi treatment, relative to phosphate-buffered saline (PBS) treatment, with the exception that TNFR2 was significantly decreased and internalized in HCT-5 cells by ETN treatment. The HTLV-I associated genes Tax and HBZ and the PVL levels were not significantly changed. Immunofluorescence staining of HCT-5 for an HTLV-I-associated protein, GAG, was also not significantly different between any of the TNFi treatments and the PBS treatment. DNA ladders as an index of apoptosis were not detected. Apoptotic cells were not increased by the addition of any TNFi. CONCLUSIONS: In vitro, TNFi did not affect the cytokine profiles, expression of associated genes and proteins, proviral load or apoptosis of HCT-5 cells. The results suggested that TNFi treatment of RA patients complicated with HTLV-I might have no effect on HTLV-I infection.

Proviral Features of Human T Cell Leukemia Virus Type 1 in Carriers with Indeterminate Western Blot Analysis Results.

Western blotting (WB) for human T cell leukemia virus type 1 (HTLV-1) is performed to confirm anti-HTLV-1 antibodies detected at the initial screening of blood donors and in pregnant women. However, the frequent occurrence of indeterminate results is a problem with this test. We therefore assessed the cause of indeterminate WB results by analyzing HTLV-1 provirus genomic sequences. A quantitative PCR assay measuring HTLV-1 provirus in WB-indeterminate samples revealed that the median proviral load was approximately 100-fold lower than that of WB-positive samples (0.01 versus 0.71 copy/100 cells). Phylogenic analysis of the complete HTLV-1 genomes of WB-indeterminate samples did not identify any specific phylogenetic groups. When we analyzed the nucleotide changes in 19 HTLV-1 isolates from WB-indeterminate samples, we identified 135 single nucleotide substitutions, composed of four types, G to A (29%), C to T (19%), T to C (19%), and A to G (16%). In the most frequent G-to-A substitution, 64% occurred at GG dinucleotides, indicating that APOBEC3G is responsible for mutagenesis in WB-indeterminate samples. Moreover, interestingly, five WB-indeterminate isolates had nonsense mutations in Pol and/or Tax, Env, p12, and p30. These findings suggest that WB-indeterminate carriers have low production of viral antigens because of a combination of a low proviral load and mutations in the provirus, which may interfere with host recognition of HTLV-1 antigens.

Extraintestinal infection of Helicobacter cinaedi induced by oral administration to Balb/c mice.

Although Helicobacter cinaedi was initially considered an opportunistic pathogen in immunocompromised patients, it was later shown to also infect immunocompetent and healthy individuals. Sporadic bacteremia due to H. cinaedi has frequently been reported; however, whether the bacterium can be translocated after passage through the intestinal mucosa remains unclear. In the present study, a preclinical small animal model that faithfully reproduces H. cinaedi infection in humans was developed. Balb/c male mice were orally inoculated with a single dose of 6.8 × 107 CFU of a human clinical H. cinaedi strain. The organism persistently colonized the intestinal tract of the mice, particularly the cecum and colon, for at least 56 days, and the bacteria were excreted in the feces. Although inoculated bacteria were recovered from the spleen, liver, kidney, lung, bladder and mesenteric lymph nodes during the first 2 weeks of bacteremia, the organism was not isolated from these organs after 4 weeks, suggesting that complement- and antibody-mediated serum sensitivity account for the relatively low frequency of systemic infection. However, H. cinaedi was isolated from the biceps femoris, triceps branchii, latissimus dorsi, and trapezius muscles beyond 2 weeks after infection and after production of specific anti-H. cinaedi IgM and IgG antibodies. The present findings suggest that experimental infection of Balb/c mice with H. cinaedi may be a useful model for further studies of H. cinaedi pathogenesis, prophylaxis or therapeutic interventions in vivo.

Evaluation of Line Immunoassay to Detect HTLV-1 Infection in an Endemic Area, Southwestern Japan; Comparison with Polymerase Chain Reaction and Western Blot.

BACKGROUND: Human T-lymphotropic virus type 1 (HTLV-1) has been recognized as a cause of adult T-cell leukemia/lymphoma, HTLV-1-associated myelopathy/tropical spastic paraparesis, and HTLV-1-associated uveitis. HTLV-1 infection is normally detected by screening for HTLV-1 antibodies, and positive samples are confirmed by Western blot (WB). However, WB fails to confirm some samples that were positive for HTLV-1 antibodies on screening. Line immunoassay (LIA) is commonly used in Europe and Brazil, but not in Japan. Therefore, we evaluated the performance of LIA as a method of confirming HTLV-1 antibodies using samples in Japan. METHODS: LIA was compared with polymerase chain reaction (PCR) and WB using 50 negative and 70 positive samples tested by chemiluminescent enzyme immunoassay (CLEIA) in Miyazaki, Japan, an HTLV-1 endemic area. LIA (INNO-LIA HTLVI/II Score) and WB (Problot HTLV-I) were performed according to the manufacturer's instructions. Real-time PCR for HTLV-1 pX region was performed using DNA derived from white blood cells. The samples that tested negative by real-time PCR were further tested by nested PCR. RESULTS: All 50 CLEIA negative samples were determined to be negative by LIA and PCR. Of the 70 positive samples, 66 tested positive by both of LIA and PCR. Three samples tested negative by LIA and PCR, and the remaining sample (PCR negative) showed non-specific staining in LIA and WB. WB showed more indeterminate results than LIA. Gp21 antibody in LIA demonstrated a high ability to discriminate between positive and negative PCR results. Furthermore, the degree of gp21 antibody reaction by LIA showed correlation with HTLV-1 proviral loads (PVLs). CONCLUSIONS: Our results indicate that LIA performs well in confirming HTLV-1 seropositivity by showing a low incidence of indeterminate results and good agreement with PCR using samples in Japan, although the number of samples tested was small. In addition, semi-quantitative antibody titer to gp21 correlated well with HTLV-1 PVLs. Further study including larger samples is necessary to determine the positioning of LIA for HTLV-1 detection in Japan.

Possible Case of Novel Spotted Fever Group Rickettsiosis in Traveler Returning to Japan from India.

A 60-year-old woman experienced fever, headache, rash, and altered vision after returning to Japan from India. Testing detected elevated antibody titers to spotted fever group rickettsia; PCR on blood yielded positive results for the rickettsial outer membrane protein A gene. We isolated a unique rickettsial agent and performed a full-genome analysis.

Severe fever with thrombocytopenia syndrome with myocardial dysfunction and encephalopathy: A case report.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease in China, Korea and Japan caused by a novel bunyavirus, SFTS virus (SFTSV). Although central nervous system manifestations are common in SFTS patients, the pathogenesis has not been elucidated; and there are few reports of myocardial dysfunction. Here we report an elderly Japanese patient with reversible myocardial dysfunction and encephalopathy. A previously healthy 65-year-old male engaged in forestry got a tick bite and developed fever and fatigue in 3 days. Three days after onset, he presented to a local hospital where the diagnosis of SFTS with hemophagocytotic syndrome was made. The blood test showed leukopenia and thrombocytopenia as well as elevated levels of alanine aminotransferase and aspartate aminotransferase. Marked hemophagocytosis was found on bone marrow smear. Peripheral blood was positive for SFTSV gene by reverse-transcription polymerase chain reaction. On day 7, the patient was transferred to our hospital. We observed disturbance of consciousness, Kernig sign and myoclonus to face and limbs. Decreased blood flow of whole cerebral cortex was detected by single photon emission computed tomography (SPECT). Chest X-ray revealed cardiomegaly and electrocardiography (ECG) showed abnormal T waves. These data suggested acute encephalopathy and myocardial dysfunction. We treated him with corticosteroid and blood transfusion, which resulted in the complete recovery of the above abnormal symptoms and laboratory data including the findings in SPECT and ECG in about a month. This case demonstrated transient myocardial dysfunction and encephalopathy can occur in addition to typical clinical manifestation of SFTS.

Standardization of Quantitative PCR for Human T-Cell Leukemia Virus Type 1 in Japan: a Collaborative Study.

Quantitative PCR (qPCR) analysis of human T-cell leukemia virus type 1 (HTLV-1) was used to assess the amount of HTLV-1 provirus DNA integrated into the genomic DNA of host blood cells. Accumulating evidence indicates that a high proviral load is one of the risk factors for the development of adult T-cell leukemia/lymphoma and HTLV-1-associated myelopathy/tropical spastic paraparesis. However, interlaboratory variability in qPCR results makes it difficult to assess the differences in reported proviral loads between laboratories. To remedy this situation, we attempted to minimize discrepancies between laboratories through standardization of HTLV-1 qPCR in a collaborative study. TL-Om1 cells that harbor the HTLV-1 provirus were serially diluted with peripheral blood mononuclear cells to prepare a candidate standard. By statistically evaluating the proviral loads of the standard and those determined using in-house qPCR methods at each laboratory, we determined the relative ratios of the measured values in the laboratories to the theoretical values of the TL-Om1 standard. The relative ratios of the laboratories ranged from 0.84 to 4.45. Next, we corrected the proviral loads of the clinical samples from HTLV-1 carriers using the relative ratio. As expected, the overall differences between the laboratories were reduced by half, from 7.4-fold to 3.8-fold on average, after applying the correction. HTLV-1 qPCR can be standardized using TL-Om1 cells as a standard and by determining the relative ratio of the measured to the theoretical standard values in each laboratory.

Use of anti-tumor necrosis factor biologics in the treatment of rheumatoid arthritis does not change human T-lymphotropic virus type 1 markers: a case series.

Anti-tumor necrosis factor (anti-TNF) biologics are effective in the treatment of rheumatoid arthritis (RA); however, it is still not clear whether this treatment promotes the development of malignancies such as lymphoma. Human T-lymphotropic virus type 1 (HTLV-1), which is a causative agent of adult T-cell lymphoma (ATL), is prevalent in Japan. Many HTLV-1-positive patients with RA are assumed to exist; however, there have thus far been no reports on the effect of anti-TNF biologics on HTLV-1-positive patients. We analyzed the response to treatment with anti-TNF biologics and change of HTLV-1 markers in two cases of RA. The two cases showed no response based on the European League Against of Rheumatism response criteria 60-96 weeks after administration of anti-TNF biologics (infliximab and etanercept). No signs of ATL were observed and HTLV-1 markers, such as proviral load and clonality of HTLV-1-infected cells, showed no significant change in either of two cases. Therefore, treatment with anti-TNF biologics did not induce activation of HTLV-1, although the effect on RA was not as effective as in HTLV-1-negative patients in this limited study. Further long-term study with a greater number of patients is necessary to clarify the safety and efficacy of anti-TNF biologics in HTLV-1-positive patients with RA.

Rapid identification and subtyping of Helicobacter cinaedi strains by intact-cell mass spectrometry profiling with the use of matrix-assisted laser desorption ionization-time of flight mass spectrometry.

Helicobacter cinaedi infection is recognized as an increasingly important emerging disease in humans. Although H. cinaedi-like strains have been isolated from a variety of animals, it is difficult to identify particular isolates due to their unusual phenotypic profiles and the limited number of biochemical tests for detecting helicobacters. Moreover, analyses of the 16S rRNA gene sequences are also limited due to the high levels of similarity among closely related helicobacters. This study was conducted to evaluate intact-cell mass spectrometry (ICMS) profiling using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) as a tool for the identification of H. cinaedi. A total of 68 strains of H. cinaedi isolated from humans, dogs, a cat, and hamsters were examined in addition to other Helicobacter species. The major ICMS profiles of H. cinaedi were identical and differed from those of Helicobacter bilis, which show >98% sequence similarity at the 16S rRNA sequence level. A phyloproteomic analysis of the H. cinaedi strains examined in this work revealed that human isolates formed a single cluster that was distinct from that of the animal isolates, with the exception of two strains from dogs. These phyloproteomic results agreed with those of the phylogenetic analysis based on the nucleotide sequences of the hsp60 gene. Because they formed a distinct cluster in both analyses, our data suggest that animal strains may not be a major source of infection in humans. In conclusion, the ICMS profiles obtained using a MALDI-TOF MS approach may be useful for the identification and subtyping of H. cinaedi.

Increased plasma lactoferrin levels in leucocytapheresis therapy in patients with rheumatoid arthritis.

OBJECTIVE: The aim of this study was to clarify the mechanism of leucocytapheresis (LCAP) in patients with RA. METHODS: Protein profiles of blood samples from two patients with RA obtained via LCAP column inlet and outlet lines were analysed by two-dimensional fluorescence difference gel electrophoresis and mass spectrometry. The lactoferrin (LTF) levels of peripheral and circulating blood samples from seven patients obtained via the LCAP column blood circuit were then determined by ELISA. Peripheral blood samples from 14 patients with RA were exposed to unwoven polyester fibre filters and the LTF level was determined. In addition, morphological changes in neutrophils after exposure to the filter were examined by optical microscopy, electronic microscopy and LTF immunostaining. RESULTS: LTF levels were increased in both samples from the LCAP column outlet and peripheral blood at the end of LCAP treatment. Furthermore, peripheral blood samples exposed to the filter revealed a decreased number of neutrophils and an increased level of LTF. Morphological analysis of the exposed neutrophils showed vacuolization of the cytoplasm and degranulation of LTF-positive granules. These data suggest that LTF stored in the granules of neutrophils is released from the neutrophils caught in the LCAP column. CONCLUSION: Because LTF has been reported to have multiple anti-inflammatory properties, increased levels of LTF may contribute to the clinical effect of LCAP in patients with RA.

Comparative evaluation of agar dilution and broth microdilution methods for antibiotic susceptibility testing of Helicobacter cinaedi.

The aim of this study was to establish a broth microdilution method for antimicrobial susceptibility testing of Helicobacter cinaedi and to assess the prevalence and mechanisms of fluoroquinolone resistance in Japanese clinical isolates. A broth microdilution method using modified Levinthal broth was developed and compared with the agar dilution method for testing susceptibility to ampicillin, gentamicin, tetracycline and ciprofloxacin. The minimum inhibitory concentrations obtained by these two methods were almost the same for all the antibiotics tested, demonstrating the broth microdilution method to be a suitable and reliable technique for antimicrobial susceptibility testing. A broth microdilution method for antimicrobial susceptibility test for H. cinaedi was established. This method is expected to help improve treatment.

Proviral loads of human T-lymphotropic virus Type 1 in asymptomatic carriers with different infection routes.

High human T-lymphotropic virus Type 1 (HTLV-1) proviral DNA load (PVL) has been reported to be one risk factor for the development of adult T-cell leukemia/lymphoma (ATL). ATL is also believed to develop in HTLV-1 carriers who acquire infection perinatally. ATL cells have been reported to frequently harbor defective provirus. In our study, PVLs for three different regions of HTLV-1 provirus (5'LTR-gag, gag and pX) were measured in 309 asymptomatic carriers with different infection routes. PVLs for the pX region in 21 asymptomatic carriers with maternal infection was significantly higher than in 24 carriers with spousal infection. Among 161 carriers with relatively high pX PVLs (equal to or greater than 1 copy per 100 peripheral blood mononuclear cells), 26 carriers (16%) had low gag PVL/pX PVL (less than 0.5) and four (2%) had low 5'LTR-gag PVL/pX PVL (less than 0.5). Low gag PVL/pX PVL ratio, which reflects deficiency and/or polymorphism of HTLV-1 proviral DNA sequences for the gag region, was also associated with maternal infection. These data suggest that HTLV-1 carriers with maternal infection tend to have high PVLs, which may be related to provirus with deficiency and/or the polymorphism of proviral DNA sequences. In addition, there is a possibility that this ratio may be used as a tool to differentiate the infection routes of asymptomatic HTLV-1 carriers, which supports the need for a large scale study.

Human T-cell leukemia virus type I (HTLV-1) proviral load and disease progression in asymptomatic HTLV-1 carriers: a nationwide prospective study in Japan.

Definitive risk factors for the development of adult T-cell leukemia (ATL) among asymptomatic human T-cell leukemia virus type I (HTLV-1) carriers remain unclear. Recently, HTLV-1 proviral loads have been evaluated as important predictors of ATL, but a few small prospective studies have been conducted. We prospectively evaluated 1218 asymptomatic HTLV-1 carriers (426 males and 792 females) who were enrolled during 2002 to 2008. The proviral load at enrollment was significantly higher in males than females (median, 2.10 vs 1.39 copies/100 peripheral blood mononuclear cells [PBMCs]; P < .001), in those 40 to 49 and 50 to 59 years of age than that of those 40 years of age and younger (P = .02 and .007, respectively), and in those with a family history of ATL than those without the history (median, 2.32 vs 1.33 copies/100 PBMCs; P = .005). During follow-up, 14 participants progressed to overt ATL. Their baseline proviral load was high (range, 4.17-28.58 copies/100 PBMCs). None developed ATL among those with a baseline proviral load lower than approximately 4 copies. Multivariate Cox analyses indicated that not only a higher proviral load, advanced age, family history of ATL, and first opportunity for HTLV-1 testing during treatment for other diseases were independent risk factors for progression of ATL.