Cytokine profiles in an extremely preterm infant with congenital syphilis.

We report the cytokine profiles of an infant with congenital syphilis as a first case. This female infant was born by vaginal delivery at a gestational age of 27 weeks during her mother's treatment for syphilis. Elevation of T helper (Th)-1 cytokines (interleukin (IL)-2, IL-12) and IL-17, which supports immunological mechanisms of Th-1, was similar to that in cases of syphilis in adults. IL-6 and granulocyte colony-stimulating factor (G-CSF), the synergistic effects of which cause the leukemoid reaction, were also elevated. The levels of interferon-γ and IL-17 in cerebral spinal fluid, which are elevated in neurosyphilis in adults, were slightly elevated.

Urinary and serum levels of high mobility group box 1.

Background: High mobility group box 1 (HMGB-1) has been extensively studied in adults and to a certain extent in neonates as well. Clinical examination of neonates, especially unwell neonates soon after birth, should be minimally invasive. Objective: This study aimed to investigate whether the urinary HMGB-1 level is comparable to the serum HMGB-1 level in neonates. Methods: In all, 87 neonates (37.5 ± 2.9 weeks of gestation and a mean birth weight of 2588 ± 649 g) were enrolled. Of these, 53 were males and 34 were females. The umbilical cord blood and the first or second spontaneous voiding urine samples were stored, and the HMGB-1 level in the samples was measured. Results: HMGB-1 was detected in all urinary samples. In these samples, we found acetylated HMGB-1 and may be devoid of nine residues at the N-terminal amino acid sequence. There was a significant correlation between the serum HMGB-1 level and urinary HMGB-1 level (r = 0.73, p < 0.001). Urinary HMGB1 levels in fetal neonatal asphyxia were significantly higher than those in healthy controls (p = 0.09). Conclusion: Urinary excretion may be one of the metabolic pathways of HMGB-1. The urinary HMGB-1 level may be comparable to the serum HMGB-1 level in the early neonatal period.

Investigation of biomarkers in a rare case of fulminant necrotizing enterocolitis in a preterm infant.

We encountered a very rare case of fulminant necrotizing enterocolitis (F-NEC) in a preterm male baby. The course of NEC and sepsis in this case was clearly different from the usual course. After onset at 14 days of life, catheter-related bloodstream infection was first assumed, and antibiotics and γ-globulin administration were started. However, 12 hours after onset, the baby's abdominal distension increased remarkably, and his entire abdominal wall turned red to purple. Escherichia coli were isolated from the blood culture, but the catheter tip culture was negative. Exchange transfusion was performed 32 hours after onset, but no significant changes were observed in the baby's general condition, and he died 46 hours after onset. The acute phase reactants of CRP and α1-acid glycoprotein increased, but haptoglobin did not. Although IL-1ß and TNFα increased as expected with sepsis, IL-6, IL-8, IL-10, and G-CSF however increased to a greater extent than expected. From the above, we diagnosed the development of intestinal necrosis as a result of widespread intestinal ischemia, and that sepsis was associated with this poor condition.

Improving water dispersibility and bioavailability of luteolin using microemulsion system.

We have studied the physiological effects and health functions of luteolin, especially focusing on its absorption and metabolism. Recent studies have reported the advantages of microemulsion to improve the bioavailability of poorly water-soluble compounds, including luteolin. In the present study, we aimed to evaluate the absorption and metabolic profile of luteolin delivered in microemulsion system via oral intake. First, we prepared water-dispersed luteolin (WD-L) using a microemulsion-based delivery system and confirmed that WD-L has superior water dispersibility compared to free luteolin (CO-L) based on their particle size distributions. Following administration of WD-L and CO-L to rats, we detected high level of luteolin-3'-O-ß-glucuronide and lower levels of luteolin, luteolin-4'-O-ß-glucuronide, and luteolin-7-O-ß-glucuronide in plasma from both CO-L and WD-L groups, indicating that the metabolic profile of luteolin was similar for both groups. On top of that, we found a 2.2-fold increase in the plasma area under the curve (AUC) of luteolin-3'-O-ß-glucuronide (main luteolin metabolite) in WD-L group (vs. CO-L). Altogether, our results suggested that delivering luteolin by microemulsion system improve its oral bioavailability without affecting its metabolite profile. This evidence thereby provides a solid basis for future application of microemulsion system for optimal delivery of luteolin.

Peripheral platelet phagocytosis in an extremely low birth weight infant:a case report.

A 768 g female neonate, born at 25 weeks' gestation, developed sepsis due to methicillin-resistant Staphylococcus epidermidis on day 14. Severe thrombocytopenia was observed, and hemophagocytic macrophages were identified in her peripheral blood smear. Cytokine profiles at the time of onset suggested that an inflammatory cytokine storm had activated lymphocytes and macrophages, leading to platelet phagocytosis. After administration of vancomycin for 14 days and immunoglobulin therapy, she improved without any complications. Considering the results of cytokine profiles, early intervention for infection may have prevented progression to hemophagocytic lymphohistiocytosis and reduced the severity of clinical symptoms.

Time-series analysis of population dynamics of the common cutworm, Spodoptera litura (Lepidoptera: Noctuidae), using an ARIMAX model.

BACKGROUND: Developing a model that adequately explains pest population dynamics based on weather-related parameters is fundamentally important for proper pest management. Autocorrelation with past occurrences should be considered when modeling the relationship between the time series of pest occurrence data and meteorological factors; however, few attempts have been made to model these factors simultaneously. In this study, we constructed an autoregressive integrated moving average with exogenous variables (ARIMAX) model to represent the occurrence of the common cutworm, Spodoptera litura (F.) (Lepidoptera: Noctuidae), a major moth pest species in Asia, using the trap catch data of S. litura recorded approximately every 5 days. The multiple meteorological measurements taken over several past periods before S. litura occurrence were included as explanatory variables to evaluate their lag effects on future occurrences. RESULTS: It was suggested that temperature had the most important effect on S. litura occurrences among other meteorological factors (i.e., humidity, wind speed, and precipitation). Especially, higher temperatures during the larval/egg stage seemed to presage a higher moth abundance. When the model was fitted using independent data that were not used for calibrating the model, the model was able to capture trends in increases in the scale of occurrence, particularly after July, when the occurrence rapidly increased. CONCLUSION: Past temperature condition, particularly during the larval and egg states, is suggested to be highly important for predicting future S. litura occurrences. The ARIMAX model proposed here will allow preventive measures to be taken, effectively safeguarding food resources against pest outbreaks. © 2022 Society of Chemical Industry.

Novel COL4A1 mutations identified in infants with congenital hemolytic anemia in association with brain malformations.

Genetic causes of undiagnosed hemolytic anemia in nineteen patients were analyzed by whole-exome sequencing, and novel COL4A1 variants were identified in four patients (21%). All patients were complicated with congenital malformations of the brain, such as porencephaly or schizencephaly. In these patients, hemolysis became less severe within 2 months after birth, and red cell transfusion was no longer required after 50 days, whereas chronic hemolysis continued.

A male preterm infant with cow's milk allergy to human milk fortifier showing only severe respiratory symptoms.

We report a male infant with a birthweight of 1,400 g at 29 weeks 2 days gestation diagnosed as having cow's milk allergy (CMA) due to human milk fortifier, who developed severe respiratory symptoms. The infant had no gastrointestinal symptoms; rather, the initial symptoms were apnea attacks and wheezing with a prolonged expiratory phase that progressed to severe ventilatory insufficiency requiring mechanical ventilation. Aggravation of his general condition, which appeared to be due to sepsis, was improved by temporary starvation and respiratory care, but he relapsed on the resumption of enteral feeding of his mother's milk with a human milk fortifier. As a result, this event was interpreted as a positive oral food challenge test. The infant resumed complete breastfeeding without the fortifier and has not relapsed since. Examination of his serial cytokine profiles from residual serum revealed that although interleukin-5 was not increased, interferon (IFN)-γ was increased, suggesting some relation between the time course of IFN-γ and the infant's eosinophil count. These findings may indicate that the involvement of IFN-γ is one cause of the onset of this disease.

Neonatal Leukemoid Reaction with Fetal Inflammatory Response Syndrome Is Associated with Elevated Serum Granulocyte Colony Stimulating Factor and Interleukin-6.

Leukemoid reaction (LR) is a reactive disease that exhibits abnormal blood values similar to leukemia, but not due to leukemia. One report showed that neonatal LR (NLR) was associated with elevated serum granulocyte colony stimulating factor (G-CSF) in only 30% of the study neonates. NLR is not always associated with the elevation of serum G-CSF. NLR was defined as a white blood cell count of ≥ 40 × 103/µL and/or blast cell concentration of > 2%. We have focused on NLR with fetal inflammatory response syndrome (FIRS), defined as a fetal systemic inflammatory reaction triggered by intrauterine infection. FIRS was diagnosed based on a cord serum interleukin-6 (IL-6) concentration ≥ 17.5 pg/mL and histopathological chorioamnionitis. Because NLR is highly associated with FIRS, we have hypothesized that NLR is associated with the elevation of both G-CSF and IL-6. This is the first report to measure multiple cytokines in NLR at the same time. The study comprised 19 preterm infants with FIRS: 8 with NLR (study group) and 11 without NLR (control group). Serum G-CSF and IL-6 concentrations were significantly higher in the study group than the control group. There was a positive correlation between G-CSF and IL-6 levels in the study group but not in the control group. These results suggest that elevated serum G-CSF and IL-6 may underlie NLR. Thus, G-CSF and IL-6 concentrations may be predictive of the onset of NLR. Measuring these cytokines is useful for judging the prognosis of preterm infants and for their post-natal clinical management.

Serial investigation of continuous glucose monitoring in a very low birth weight infant with transient late-onset hyperglycemia.

Transient late-onset hyperglycemia was detected in a very low birth weight (VLBW) infant (gestational age 28 weeks, birth weight 1,082 g) by routine point-of-care glucose monitoring.　The infant had no clinical symptom.　Serial continuous glucose monitoring (CGM) was conducted for 3 days at 31, 35, and 39 weeks' post conceptual age.　The difference values between the maximum and minimum blood glucose levels during the interval from one enteral feeding to the next enteral feeding were 32.3±14.3 mg/dL, 47.5±22.9 mg/dL, and 27.5±12.9 mg/dL for the 1st, 2nd, and 3rd CGM, respectively.　The serial change in the values was statistically significant (p<0.01).CGM is widely used as a routine clinical practice, which is true even in VLBW infants.　Hyperglycemic events detected by only once of CGM in otherwise healthy preterm infants have already been reported on larger numbers of patients.　To our knowledge, this is the first report that the change of glucose intolerance in a VLBW infant with transient late-onset hyperglycemia was investigated by serial CGM.

DOWN'S SYNDROME WITH NEONATAL ALLOIMMUNE THROMBOCYTOPENIA DUE TO HLA-A2 ANTIBODY.

Anti-HLA antibodies reportedly exist in the one third of pregnant women. But few occurrences of neonatal alloimmune thrombocytopenia (NAIT) caused by anti-HLA antibodies have been reported. Here a male baby, who was admitted for low birth weight with Down syndrome (DS), was suffered from thrombocytopenia without transient myeloproliferative disorder (TMD). Positive reactions of HLA-specific antibodies were detected in maternal serum. Cross-matching tests between maternal serum and paternal platelets and lymphocytes were strongly positive. It is most conceivable that the previous pregnancy of the mother induced the production of anti-HLA-A2 antibody, which crossed the placenta and subsequently caused an NAIT in the case presented. This is the first case of DS with NAIT due to anti-HLA antibodies.

Establishment of tracking system for West Nile virus entry and evidence of microtubule involvement in particle transport.

West Nile virus (WNV) is one of flaviviruses and has emerged recently in the United States as a significant cause of viral encephalitis. Although cellular entry of WNV is important for viral pathogenesis, its mechanisms have not been elucidated fully. To explore the entry mechanisms, a virus-particle tracking system in live cells by using fluorescently labeled subviral particles (SVPs) and time-lapse epifluorescence microscopy was established. This study revealed that, following cellular entry, SVP movements could be divided into two phases: early (slow movement) and late (fast movement) phase. Moreover, fast viral particle movements at the late phase correlated with SVP-microtubule association.

Purification and concentration of non-infectious West Nile virus-like particles and infectious virions using a pseudo-affinity Cellufine Sulfate column.

Affinity column chromatography is a promising method for the purification of flavivirus particles that can supplement or potentially replace diafiltration and sucrose density centrifugation. In this study, the purification of West Nile Virus (WNV) antigens via Cellufine Sulfate column chromatography was examined. Virus-like particles (VLPs) produced by the expression of the prM and E genes were separated from most of the contaminant proteins with 0.2-0.4M NaCl, but still retained their spherical forms and immunogenicity in mice. The column, with a 1 mL bed-volume, concentrated WN-VLPs a minimum of 15 fold from culture supernatants. A heparin analogue, suramin, competitively eluted WN-VLPs, but sulphated polysaccharides, such as heparin, heparin sulfate and dextran sulfate, did not. Furthermore, 2.4 × 109 plaque forming units of WNV and 196 µg of the viral antigens were recovered from 60 mL of infected culture medium at high yields (93% and 96%, respectively). These results indicate that, in addition to conventional methods, Cellufine Sulfate column chromatography is an effective preparation technique for WNV particulate antigens that does not impair the antigen virological characteristics.

Accumulation of cellular prion protein within dystrophic neurites of amyloid plaques in the Alzheimer's disease brain.

Amyloid plaques, a well-known hallmark of Alzheimer's disease (AD), are formed by aggregated ß-amyloid (Aß). The cellular prion protein (PrPc) accumulates concomitantly with Aß in amyloid plaques. One type of amyloid plaque, classified as a neuritic plaque, is composed of an amyloid core and surrounding dystrophic neurites. PrPc immunoreactivity reminiscent of dystrophic neurites is observed in neuritic plaques. Proteinase K treatment prior to immunohistochemistry removes PrPc immunoreactivity from amyloid plaques, whereas Aß immunoreactivity is enhanced by this treatment. In the present study, we used a chemical pretreatment by a sarkosyl solution (0.1% sarkosyl, 75 mM NaOH, 2% NaCl), instead of proteinase K treatment, to evaluate PrPc accumulation within amyloid plaques. Since PrPc within amyloid plaques is removed by this chemical pretreatment, we can recognize that the PrP species deposits within amyloid plaques were PrPc. We could observe that PrPc accumulation in dystrophic neurites occurred differently compared with Aß or hyperphosphorylated tau aggregation in the AD brain. These results could support the hypothesis that PrPc accumulation in dystrophic neurites reflects a response to impairments in cellular degradation, endocytosis, or transport mechanisms associated with AD rather than a non-specific cross-reactivity between PrPc and aggregated Aß or tau.

Intranasal administration of adjuvant-combined vaccine protects monkeys from challenge with the highly pathogenic influenza A H5N1 virus.

The effectiveness in cynomolgus macaques of intranasal administration of an influenza A H5N1 pre-pandemic vaccine combined with synthetic double-stranded RNA (polyI/polyC12U) as an adjuvant was examined. The monkeys were immunized with the adjuvant-combined vaccine on weeks 0, 3, and 5, and challenged with the homologous virus 2 weeks after the third immunization. After the second immunization, the immunization induced vaccine-specific salivary IgA and serum IgG antibodies, as detected by ELISA. The serum IgG antibodies present 2 weeks after the third immunization not only had high neutralizing activity against the homologous virus, they also neutralized significantly heterologous influenza A H5N1 viruses. The vaccinated animals were protected completely from the challenge infection with the homologous virus. These results suggest that intranasal immunization with the Double stranded RNA-combined influenza A H5N1 vaccine induce mucosal IgA and serum IgG antibodies which could protect humans from homologous influenza A H5N1 viruses which have a pandemic potential.

Immunogenicity and efficacy of two types of West Nile virus-like particles different in size and maturation as a second-generation vaccine candidate.

Virus-like particles (VLPs) of flaviviruses generated from the prM and E genes are a promising vaccine candidate. We have established cell clones continuously releasing VLPs of West Nile virus (WNV) in serum-free conditions. Two types of VLPs were distinguished by sedimenting analyses in sucrose density gradients. Fast sedimenting VLPs (F-VLPs) were large (40-50 nm) and composed of the E and processed mature M proteins, whereas slowly sedimenting VLPs (S-VLPs) were small (20-30 nm) particles consisting of the E and immature prM proteins. F-VLPs induced higher neutralizing antibody and anti-WNV IgG titers than S-VLPs. Furthermore, IgG2a was dominant over IgG1 by immunization with F-VLPs as with whole virion-derived antigens. Mice vaccinated with a low dose (3 ng) of F-VLPs showed higher protective efficacy (83% survivals) against WNV infection than S-VLP-immune mice (17% survivals). These results indicate that F-VLPs more closely resemble the virions and take a better immunogenic form than S-VLPs as WNV vaccine candidates.

ImmunoAT method: An initial assessment for the detection of abnormal isoforms of prion protein in formalin-fixed and paraffin-embedded tissues.

The AT-tailing method is a labelling technique that utilises oligo(dA-dT)-dependent signal amplification. In this study, a new immunohistochemical application of the immunoAT method was developed. This method uses an oligo(dA-dT)-conjugated primary antibody (direct immunoAT method) or an oligo(dA-dT)-conjugated secondary antibody (indirect immunoAT method). Fifteen-base oligo(dA-dT)-conjugated antibodies (IgG-ATs) were prepared in advance by conjugating maleimide-activated oligo(dA-dT) to IgG via free sulfhydryl residues that had been introduced on the surface of IgG using Traut's reagent. Following the reaction with the target antigen and the IgG-AT, oligo(dA-dT) was elongated by DeltaTth DNA polymerase in the presence of dATP, dTTP and biotinylated dUTP, consequently labelling the antigen-antibody complex with a large amount of biotin. To initially evaluate the immunoAT method, the presence or absence of prion protein (PrP(sc)) was determined in formalin-fixed and paraffin-embedded sections of the medulla oblongata of cattle which had been under active surveillance for bovine spongiform encephalopathy. Sections were examined using direct and indirect immunoAT methods and the EnVision+ system (Dako) under conditions that were identical except for the differing IgG-AT and AT-tailing methods. PrP(sc) detection was consistent using all three methods. The clearest signals were obtained using the indirect immunoAT method, suggesting significant potential for this method.

Induction of cross-protective immunity against influenza A virus H5N1 by an intranasal vaccine with extracts of mushroom mycelia.

The identification of a safe and effective adjuvant that is able to enhance mucosal immune responses is necessary for the development of an efficient inactivated intranasal influenza vaccine. The present study demonstrated the effectiveness of extracts of mycelia derived from edible mushrooms as adjuvants for intranasal influenza vaccine. The adjuvant effect of extracts of mycelia was examined by intranasal co-administration of the extracts and inactivated A/PR8 (H1N1) influenza virus hemagglutinin (HA) vaccine in BALB/c mice. The inactivated vaccine in combination with mycelial extracts induced a high anti-A/PR8 HA-specific IgA and IgG response in nasal washings and serum, respectively. Virus-specific cytotoxic T-lymphocyte responses were also induced by administration of the vaccine with extract of mycelia, resulting in protection against lethal lung infection with influenza virus A/PR8. In addition, intranasal administration of NIBRG14 vaccine derived from the influenza A/Vietnam/1194/2004 (H5N1) virus strain administered in conjunction with mycelial extracts from Phellinus linteus conferred cross-protection against heterologous influenza A/Indonesia/6/2005 virus challenge in the nasal infection model. In addition, mycelial extracts induced proinflammatory cytokines and CD40 expression in bone marrow-derived dendritic cells. These results suggest that mycelial extract-adjuvanted vaccines can confer cross-protection against variant H5N1 influenza viruses. The use of extracts of mycelia derived from edible mushrooms is proposed as a new safe and effective mucosal adjuvant for use for nasal vaccination against influenza virus infection.