Prognostic significance of absolute lymphocyte count in patients with metastatic renal cell carcinoma receiving first-line combination immunotherapies: results from the International Metastatic Renal Cell Carcinoma Database Consortium.

BACKGROUND: Lymphocytes are closely linked to mechanisms of action of immuno-oncology (IO) agents. We aimed to assess the prognostic significance of absolute lymphocyte count (ALC) in patients with metastatic renal cell carcinoma (mRCC). PATIENTS AND METHODS: Using the International mRCC Database Consortium (IMDC), patients receiving first-line IO-based combination therapy were analysed. Baseline patient characteristics, objective response rates (ORRs), time to next treatment (TTNT), and overall survival (OS) were compared. RESULTS: Of 966 patients included, 195 (20%) had lymphopenia at baseline, and they had a lower ORR (37% versus 45%; P < 0.001), shorter TTNT (10.1 months versus 24.3 months; P < 0.001), and shorter OS (30.4 months versus 48.2 months; P < 0.001). Among 125 patients with lymphopenia at baseline, 52 (42%) experienced ALC recovery at 3 months, and they had longer OS (not reached versus 30.4 months; P = 0.012). On multivariable analysis for OS, lymphopenia was an independent adverse prognostic factor (hazard ratio 1.68; P < 0.001). Incorporation of lymphopenia into the IMDC criteria improved OS prediction accuracy (C-index from 0.688 to 0.707). CONCLUSIONS: Lymphopenia was observed in one-fifth of treatment-naive patients with mRCC and may serve as an indicator of unfavourable oncologic outcomes in the contemporary IO era.

Conceptual design of heavy ion beam probes on the PLATO tokamak.

Heavy ion beam probe (HIBP) systems have been designed for the new tokamak, PLATO [A. Fujisawa, AIP Conf. Proc. 1993, 020011 (2018)]. The designs have been completed, and the installations are in progress. Two HIBPs are being installed in toroidal sections 180° apart to investigate long-range correlations in the toroidal direction. Each HIBP consists of an injection beamline and a detection beamline as usual. Yet, one of the HIBPs is equipped with an additional detection beamline; the measurement positions of its two detection beamlines can be placed on almost the same magnetic surface yet at poloidal angles that differ by ∼180°. The use of three detection beamlines allows us to investigate spatial asymmetry and long-range correlations in both the toroidal and poloidal directions, simultaneously. The detected beam intensity is expected to be enough for turbulence measurements in almost the entire plasma region when the electron density is up to 1 × 1019 m-3 by selecting appropriate ion species for the probe beam. Each detector has three channels 10 mm apart, allowing measurement of local structures of micro-scale turbulence. Therefore, using the HIBPs on the PLATO tokamak will enable both local and global properties of plasma turbulence to be investigated, simultaneously.

The IL15 +96522 A>T functional polymorphism is related to the differentiation of Th17 cells and the severity of Hashimoto's disease.

To clarify the association between the genetic producibility of IL-15, a pro-inflammatory cytokine, and the pathogenesis of autoimmune thyroid diseases (AITDs), we genotyped +96522 A>T and +82889 A>G polymorphisms in the IL15 gene using 127 patients with Hashimoto's disease (HD), including 55 patients with severe HD and 48 patients with mild HD; 130 patients with Graves' disease (GD), including 52 patients with intractable GD and 44 patients with GD in remission; and 79 healthy volunteers. Both the IL15 +96522 A allele and AA genotype were more frequent in patients with severe HD than in those with mild HD. The serum levels of IL-15 were higher in individuals with the IL15 +96522 AA genotype than in those with the T allele, and they were also higher in patients with severe HD than in those with mild HD. On the other hand, the mRNA levels of IL-15 were not significantly different among individuals with each genotype of both SNPs. After incubation with recombinant human IL-15, the proportions of Th17 cells in CD4+ cells were increased, and those of Treg cells in CD4+ cells were maintained. Our study indicates that the IL15 +96522A/C polymorphism correlates with the severity of HD, most likely by increasing Th17 cells.

Eritoran inhibits S100A8-mediated TLR4/MD-2 activation and tumor growth by changing the immune microenvironment.

S100A8/A9 is a major component of the acute phase of inflammation, and appears to regulate cell proliferation, redox regulation and chemotaxis. We previously reported that S100A8/S100A9 are upregulated in the premetastatic lung. However, the detailed mechanisms by which S100A8 contributes to tumor progression have not been elucidated. In this study, we investigated the TLR4/MD-2 dependency by S100A8 on tumor progression. We found that S100A8 (2-89) peptide stimulated cell migration in a manner dependent on TLR4, MD-2 and MyD88. The S100A8 (2-89) peptide also activated p38 and NF-κB in TLR4-dependent manner. The peptide induced the upregulation of both IL-6 and Ccl2 in peritoneal macrophages obtained from wild-type mice, but not TLR4-deficient mice. We then investigated the responsible region of S100A8 for TLR4/MD-2 binding by a binding assay, and found that C-terminal region of S100A8 binds to TLR4/MD-2 complex. To further evaluate the TLR4 dependency on tumor microenvironment, Lewis lung carcinoma-bearing mice were treated with Eritoran, an antagonist of TLR4/MD-2 complex. We found that both tumor volume and pulmonary recruitment of myeloid-derived suppressor cells were reduced with the treatment of Eritoran for five consecutive days. Eritoran reduced the development of tumor vasculature, and increased tumor-infiltration of CD8(+) T-cells. Taken together, S100A8 appears to play a crucial role in the activation of the TLR4/MD-2 pathway and the promotion of a tumor growth-enhancing immune microenvironment.

Effective cell collection method using collagenase and ultrasonic vibration.

This study proposes a novel cell collection method based on collagenase treatment and ultrasonic vibration. The method collects calf chondrocytes from a reusable metal cell culture substrate. To develop our concept, we calculated the natural vibration modes of the cell culture substrate by a finite element method, and conducted eigenvalue and piezoelectric-structural analyses. Selecting the first out-of-plane vibration mode of the substrate, which has a single nodal circle, we designed and fabricated the cell collection device. The excited vibration mode properly realized our intentions. We then evaluated the cell collection ratio and the growth response, and observed the morphology of the collected cells. The collagenase and ultrasonic vibration treatment collected comparable numbers of cells to conventional trypsin and pipetting treatment, but improved the proliferating cell statistics. Morphological observations revealed that the membranes of cells collected by the proposed method remain intact; consequently, the cells are larger and rougher than cells collected by the conventional method. Therefore, we present a promising cell collection method for adhesive cell culturing process.

A triploid partial mole placenta from paternal isodisomy with a diploid fetus derived from one sperm and one oocyte may have caused angiogenic imbalance leading to preeclampsia-like symptoms at 19 weeks of gestation.

Partial hydatidiform mole with a normal fetus is extremely rare. A 30-year-old woman presented at 19 weeks gestation with clinical manifestations of severe preeclampsia. The fetus revealed a normal 46,XX karyotype and the placenta revealed triploid 69,XXX from paternal isodisomy. Microsatellite analysis revealed that the fetus and the triploid partial mole were derived from one sperm and one oocyte, followed by duplication of paternal chromosomes in only a trophectodermal cell. The maternal serum levels of angiogenic factors were extremely high compared with those reported in preeclampsia, suggesting an angiogenic imbalance may have caused preeclampsia-like symptoms before 20 weeks of gestation.

Freezing of glycerol-water mixtures under pressure.

We investigated freezing of pure glycerol as well as glycerol-water (GW) mixtures with 3:1 and 3:2 volume fractions as a function of pressure in the 0-10 GPa range by ruby fluorescence spectroscopy and neutron scattering. We find that the glass transition pressure increases from 5.5 GPa for pure glycerol to 6.5 GPa for the 3:1 GW mixture, with unusually small pressure gradients above. For higher water concentrations close to 3:2, phase separation occurs above 2 GPa where most of the water is expelled in the form of ice VII. The results suggest that glycerol is able to effectively hydrogen bond not more than ≈2.5 H(2)O molecules per glycerol, which seems to support conclusions from molecular dynamics simulations. The data indicate that these fluids could become important as pressure transmitting media for neutron scattering in the 0-7 GPa range, including at low temperatures.

Association of functional GITR gene polymorphisms related to expression of glucocorticoid-induced tumour necrosis factor-receptor (GITR) molecules with prognosis of autoimmune thyroid disease.

The glucocorticoid-induced tumour necrosis factor (TNF)-receptor (GITR) affects the functions of regulatory T (T(reg)) and effector T (T(eff)) cells, but the significance of this phenomenon is still unclear. To examine the association of single nucleotide polymorphisms (SNPs) in the GITR gene with the expression of GITR molecules on T cells and with the pathological conditions in patients with autoimmune thyroid disease (AITD), we examined the frequencies of four candidate SNPs in AITD patients and healthy volunteers by restriction enzyme analysis and direct sequence analyses. We also analysed the GITR expression on peripheral T(reg) and T(eff) cells in AITD patients by three-colour flow cytometry. The CC genotype in the rs3753348 C/G SNP was significantly more frequent in patients with mild Hashimoto's disease (HD) than in those with severe HD [P = 0·0117, odds ratio (OR) = 3·13]. The AA genotype in the rs2298213 A/G SNP was significantly more frequent in patients with mild HD than in patients with severe HD (P = 0·010, OR = 4·43). All patients and healthy individuals had the GG genotype in rs60038293 A/G and rs11466696 A/G SNPs. The proportions of GITR(+) cells in T(reg) and T(eff) cells were significantly higher in AITD patients with the CC genotype of the rs3753348 SNP than in those with the GG genotype (P = 0·004 and P = 0·011, respectively). In conclusion, the rs3753348 C/G SNP in the GITR is associated with HD prognosis and expression on T(reg) and T(eff) cells.

Knowledge and impressions regarding the concept of mutation among Japanese university students.

Although the term mutation is frequently used in genetic counseling, it may carry negative connotations and create misunderstanding. Our objective was to investigate the relationship between the impressions regarding three Japanese terms mutation of gene, change of gene, and lesion of gene as well as to investigate the depth of understanding regarding mutation. A total of 175 university students and auditing students were included and responded to two questionnaires that were Impressions regarding the term in the semantic differential method and Knowledge about the concept of mutation. In factor analysis, three factors (Value, Change Rate, and Intention) were extracted. Participants were divided into three groups depending on their knowledge, and a two-way analysis of variance (Term x Knowledge Group) was conducted on the factor score for each. Results showed that the main effect of the 'Term' was significant for the Value Factor and that interaction was significant for the Change Rate Factor, and that the main effect of Knowledge Group was significant for the Intention Factor. The findings suggest that healthcare professionals should demonstrate an awareness of varying impressions of the different terms used to refer to the identical concepts of mutation. This is of particular importance when communicating with patients and their families.

Predictive assay of neoadjuvant chemotherapy in management of oral cancer.

Neoadjuvant chemotherapy (NAC) for oral squamous cell carcinoma has a positive impact on organ preservation and/or survival only in patients who achieve an excellent anti-tumour effect with this therapy. Predictive assay for NAC can play an important role in establishing tailor-made treatments for oral squamous cell carcinoma. In this retrospective study, the anti-tumour effects of cisplatin-based NAC in 70 patients with oral squamous cell carcinoma were reviewed in relation to biological markers of tumour cell proliferation activity: tumour grade, cellular DNA content, mitotic index, apoptotic index, ki-67 positive rate, and p53 and Bax expression. Tumour grade, Bax expression, apoptotic index and cellular DNA content were significantly correlated with the anti-tumour effects of NAC in univariate analysis. Tumour grade, Bax expression and apoptotic index were selected as independent predictive factors by means of multiple logistic analysis. Using the regression equation from these results, the prediction rate for anti-tumour effects was 70%. For patients in whom NAC is predicted to be ineffective, it may be necessary to choose another treatment option in order to improve their survival and quality of life.

Evaluation of MBR effluent characteristics for reuse purposes.

One of the advantages of MBR is its excellent effluent quality, which is suitable for a wide range of reuse purposes. We investigated the characteristics of MBR effluent and evaluated them based on the Japanese guideline for the reuse of treated wastewater. As the result, MBR effluent showed qualitative coaracteristics that satisfy the requirement except chromaticity for recreational purpose. Further treatment, such as by oxone or activated carbon, will be required to remove the remaining color. MBR shows high removal efficiency of bacteria and other hazardous microorganisms such as Cryptosporidium. We investigated the removal efficiency of virus by MBR using coliphage as an alternative index. The results showed that high removal efficiency for coliphage could be obtained by MBR. The removal mechanism appears to be that coliphage are attached to the activated sludge and thus rejected by the membrane together with activated sludge particles. With regard to the endocrine disrupters, no significant differences were observed between MBR and CAS in the removal of main endocrine disrupters.

Growth responses of Escherichia coli to immunoglobulin G from cows immunized with ferric citrate receptor, FecA.

Effects of purified immunoglobulin (Ig) G from cows immunized with ferric citrate receptor, FecA, on the in vitro growth of Escherichia coli were investigated. Twenty-one cows were assigned to one of 3 treatments: 1) FecA immunization, 2) E. coli J5 bacterin immunization, and 3) unimmunized control. FecA was derived from E. coli UT5600/pSV66. Immunoglobulin G was purified from pooled colostral whey for each treatment group. The IgG from FecA immunized cows had higher titers against FecA compared with other treatment groups. Bacterial isolates tested were 14 E. coli from intramammary infections and E. coli UT5600/pSV66. Iron depletion decreased the growth of E. coli compared with growth in Fe-replete medium. The presence of IgG further decreased the growth compared with the growth under iron restriction alone. Bacterial growth did not differ among IgG sources nor between IgG concentrations. Replenishing media with exogenous iron overrode the inhibitory effects of the Fe-depletion and IgG. Vaccinating cows with FecA had little effect on the growth inhibitory properties of IgG toward E. coli mastitis isolates cultured in Fe-deplete media.

An overview of transmissible spongiform encephalopathies.

Transmissible spongiform encephalopathies (TSEs) are fatal neurodegenerative disorders of humans and animals associated with an accumulation of abnormal isoforms of prion protein (PrP) in nerve cells. The pathogenesis of TSEs involves conformational conversions of normal cellular PrP (PrP(c)) to abnormal isoforms of PrP (PrP(Sc)). While the protein-only hypothesis has been widely accepted as a causal mechanism of prion diseases, evidence from more recent research suggests a possible involvement of other cellular component(s) or as yet undefined infectious agent(s) in PrP pathogenesis. Although the underlying mechanisms of PrP strain variation and the determinants of interspecies transmissibility have not been fully elucidated, biochemical and molecular findings indicate that bovine spongiform encephalopathy in cattle and new-variant Creutzfeldt-Jakob disease in humans are caused by indistinguishable etiological agent(s). Cumulative evidence suggests that there may be risks of humans acquiring TSEs via a variety of exposures to infected material. The development of highly precise ligands is warranted to detect and differentiate strains, allelic variants and infectious isoforms of these PrPs. This article describes the general features of TSEs and PrP, the current understanding of their pathogenesis, recent advances in prion disease diagnostics, and PrP inactivation.

Effect of immunoglobulin G from cows immunized with ferric citrate receptor (FecA) on iron uptake by Escherichia coli.

The effects of immunoglobulin (Ig) G from cows immunized with the ferric citrate receptor (FecA) on iron uptake by Escherichia coli were investigated. Receptor FecA was purified from E. coli UT5600/pSV66. Cows were immunized with 400 microg purified FecA three times at 21 d intervals during late lactation and the nonlactating period. Immunoglobulin G was purified by protein G affinity chromatography from colostral whey from cows immunized with FecA and from unimmunized control cows. The purified IgG from FecA immunized cows had higher IgG titers against FecA compared with control IgG. Fifteen E. coli isolated from intramammary infections and E. coli UT5600/pSV66 were grown in an iron-depleted medium containing 1 mM citrate to induce FecA. The bacterial cells were mixed with 0, 2, and 4 mg/ml purified IgG, and 55Fe was added to the assay. After 5, 10, and 15 min incubations at 37 degrees C, samples were passed through 0.45-pm pore size filters. Filters were washed with saline three times, and the radioactivity of 55Fe taken up by the bacterial cells on the filters was measured by a liquid scintillation counter. The measurements were expressed as numbers of 55Fe atoms per colony-forming unit and transformed to log10. The assay was repeated three times for each isolate in a partially balanced incomplete block design. The presence of IgG decreased 55Fe uptake by E. coli mastitis isolates and E. coli UT5600/pSV66. Anti-FecA IgG reduced 55Fe uptake by E. coli greater than IgG from unimmunized cows.

Opsonic activity of serum and whey from cows immunized with the ferric citrate receptor.

The effects of immunizing dairy cows with the ferric citrate receptor, FecA, on the opsonic activity of serum and whey were measured in a phagocytosis assay. Fifteen cows were assigned to five blocks of three cows based on date of expected parturition. Cows within a block were randomly assigned to one of three treatments: 1) FecA immunization, 2) immunization with a commercially available Escherichia coli J5 bacterin, and 3) unimmunized controls. Cows were challenged at approximately 21 DIM by intramammary infusion of E. coli 727 into one mammary quarter. Escherichia coli 727 were opsonized for the phagocytosis assay with either 10% heat-inactivated serum or 50% heat-inactivated whey collected from each cow at calving, immediately before challenge and 7 d after challenge. Cows immunized with FecA or the E. coli J5 bacterin had increased IgG titers against FecA and E. coli 727 compared with unimmunized control cows. However, sera and whey collected from cows immunized with FecA did not enhance opsonization of E. coli 727 compared with sera and whey from control cows. Immunization with the E. coli J5 bacterin increased opsonization of sera greater than immunization with FecA. Immunoglobulin M antibody titer against E. coli 727 in whey and phagocytic indexes were positively correlated. The phagocytic index of whey immediately before challenge and 7 d after challenge were negatively associated with peak bacterial counts in mammary quarters challenged with E. coli 727. Results of the current trial suggest that the immune response resulting from immunization with FecA did not enhance opsonization and in vitro phagocytosis of E. coli 727.

Efficacy of immunization with ferric citrate receptor FecA from Escherichia coli on induced coliform mastitis.

The effects of immunization with the ferric citrate receptor FecA on antibody responses and on experimentally induced mastitis following intramammary challenge were investigated. Twenty-one cows were assigned to seven blocks of three cows based on expected parturition. Cows within block were randomly assigned to one of three treatments: 1) FecA immunization, 2) Escherichia coli J5 immunization, and 3) unimmunized controls. Challenge was by infusion of approximately 60 cfu of E. coli 727 into one uninfected mammary gland between 13 and 31 d after parturition. Cows within block were challenged on the same day. Cows immunized with FecA had higher immunoglobulin (Ig)G titers against FecA in serum and in mammary secretions at calving, immediately before challenge, and 7 d after challenge than did cows immunized with E. coli J5 or control cows. Immunization with FecA also increased IgG titers against whole-cell E. coli 727 in serum and in mammary secretions at calving. Serum IgM titers against FecA were higher in FecA immunized cows than in other treatment groups immediately before challenge. Bacterial counts in milk, duration of bacterial isolation in milk, rectal temperature, and milk somatic cell counts following intramammary challenge were similar among treatments. Milk production and dry matter intake did not differ among treatments. The ferric citrate receptor FecA was immunogenic in cows, but immunization had minimal effect on the clinical severity of experimentally induced E. coli mastitis.

Detection of cyclin D1 overexpression by real-time reverse-transcriptase-mediated quantitative polymerase chain reaction for the diagnosis of mantle cell lymphoma.

The diagnosis of mantle cell lymphoma (MCL) is particularly important for clinical management because of a remarkable prognostic difference between MCL and other types of B-cell lymphoma. In addition to immunohistochemical analysis, we have established a 5' exonuclease-based real-time reverse transcriptase-mediated quantitative polymerase chain reaction (RQ-PCR) method to detect cyclin D1 overexpression for the diagnosis of MCL. The RQ-PCR could detect cyclin D1 overexpression in all nine examined MCL cases, in contrast genomic PCR detected t(11;14) in only two of nine cases. By RQ-PCR the expression of G6PDH was significantly higher in myeloid leukemias than those in B-cell lymphomas (P = 0.018). As a result, cyclin D1/G6PDH ratio ranged from 0.78 to 12.4 (mean, 1.83) in MCL, exclusively higher than those in other B-cell lymphoma (0.00009 approximately 0.16) and myeloid leukemia (0.00011 approximately 0.085). The high expression of cyclin D1 in certain myeloid leukemias was identified to reflect their proliferative activity and not to represent the oncogenic overexpression. The 95% confidence interval of the cyclin D1/G6PDH ratio was 0.29 approximately 11.1 for MCL, 0.014 approximately 0.25 for other B-cell lymphomas and 0.000014 approximately 0.083 for myeloid leukemia, suggesting that a cutoff value can be set at 0.25. The RQ-PCR of cyclin D1 is convenient and especially useful for the diagnosis of MCL.

Immunohistochemical localization of Betacellulin, a member of epidermal growth factor family, in atherosclerotic plaques of human aorta.

Betacellulin (BTC), a new member of the EGF family, has been reported to be a potent mitogen for rat vascular smooth muscle cells (SMCs). BTC mRNA is known to be expressed in several human organs. However, the localization of BTC in human vascular tissues has not yet been clarified. We investigated whether or not BTC protein is involved in the pathogenesis of human atherosclerosis. Recombinant human BTC showed a mitogenic activity on cultured human aortic SMCs by measuring [3H]thymidine incorporation. The immunohistochemical localization of BTC, SMCs, macrophages, EGF receptors and ErbB4 was examined in autopsied human aortas. BTC was detected in both intimal and medial SMCs of the aortic wall. The percentage of BTC-positive medial SMCs in early types of atherosclerotic lesions decreased with age, but in adult, it was significantly higher in advanced types than in early types of atherosclerotic lesions. BTC-positive SMCs were predominantly localized in the medial side of the intima. Furthermore, numerous BTC-positive SMCs and macrophages were observed around the core lesion of atherosclerotic plaques. Receptors for BTC, EGF receptor and ErbB4, were expressed on SMCs, suggesting that BTC is associated with EGF receptor family-mediated signaling. BTC is produced in human aortic tissue and might play important roles in atherogenesis.

Role of nitric oxide in the synthesis of guanidinosuccinic acid, an activator of the N-methyl-D-aspartate receptor.

BACKGROUND: We propose that reactive oxygen and argininosuccinic acid (ASA) form guanidinosuccinic acid (GSA). An alternative to this hypothesis is the so-called guanidine cycle, which consists of a series of hydroxyurea derivatives that serve as intermediates in a pathway leading from urea to GSA. We compare the role of the guanidine cycle to that of nitric oxide (NO) in the synthesis of GSA. METHODS: The members of the guanidine cycle (hydroxyurea, hydroxylamine plus homoserine, L-canaline, and L-canavanine) were incubated with isolated rat hepatocytes. The known NO donors, NOR-2, NOC-7, and SIN-1, were incubated with ASA in vitro. Ornithine, arginine, or citrulline, which increase arginine, a precursor of NO, were incubated with isolated rat hepatocytes. GSA was determined by high-performance liquid chromatography. RESULTS: None of guanidine cycle members except for urea formed GSA. SIN-1, which generates superoxide and NO formed GSA, but other simple NO donors, did not. Both carboxy-PTIO, a scavenger of NO, and dimethyl sulfoxide, a hydroxyl radical scavenger, completely inhibited GSA synthesis by SIN-1. GSA formation by SIN-1 reached a maximum at 0.5 mmol/L and decreased at higher concentrations. GSA synthesis, stimulated by urea in isolated hepatocytes, was inhibited by ornithine, arginine, or citrulline with ammonia, but not by ornithine without ammonia, where arginine production is limited. CONCLUSION: GSA is formed from ASA and the hydroxyl radical. When arginine increased in hepatocytes, GSA synthesis decreased. These data suggest that increased NO, which results from high concentrations of arginine, or SIN-1 scavenges the hydroxyl radical. This may explain the decreased GSA synthesis in inborn errors of the urea cycle where ASA is decreased, and also the diminished GSA excretion in arginemia.