SARS-CoV-2 sublingual vaccine with RBD antigen and poly(I:C) adjuvant: Preclinical study in cynomolgus macaques.

Mucosal vaccine for sublingual route was prepared with recombinant SARS-CoV-2 spike protein receptor binding domain (RBD) antigen and poly(I:C) adjuvant components. The efficacy of this sublingual vaccine was examined using Cynomolgus macaques. Nine of the macaque monkeys were divided into three groups of three animals: control [just 400 µg poly(I:C) per head], low dose [30 µg RBD and 400 µg poly(I:C) per head], and high dose [150 µg RBD and 400 µg poly(I:C) per head], respectively. N-acetylcysteine (NAC), a mild reducing agent losing mucin barrier, was used to enhance vaccine delivery to mucosal immune cells. RBD-specific IgA antibody secreted in pituita was detected in two of three monkeys of the high dose group and one of three animals of the low dose group. RBD-specific IgG and/or IgA antibodies in plasma were also detected in these monkeys. These indicated that the sublingual vaccine stimulated mucosal immune response to produce antigen-specific secretory IgA antibodies in pituita and/or saliva. This sublingual vaccine also affected systemic immune response to produce IgG (IgA) in plasma. Little RBD-specific IgE was detected in plasma, suggesting no allergic antigenicity of this sublingual vaccine. Thus, SARS-CoV-2 sublingual vaccine consisting of poly(I:C) adjuvant showed reasonable efficacy in a non-human primate model.

Incidental Sacral Meningocele in an Elderly Patient Diagnosed After Epidermal Inclusion Cyst Removal: A Case Report.

A meningocele is a congenital neural tube defect, and the majority of the meningocele cases are identified perinatally. We present the case of a 67-year-old patient with a sacral meningocele undiagnosed until the removal of a symptomatic epidermal inclusion cyst adjacent to it. Cerebrospinal fluid leakage occurred due to an incision in an undiagnosed meningocele adjacent to the epidermal inclusion cyst. Repair of the cerebrospinal fluid leakage was performed successfully without any deficit. The present case underscores the importance of considering a meningocele as a differential diagnosis for a mass occurring in the midline of the back at any age.

Prevalence of saliva immunoglobulin A antibodies reactive with severe acute respiratory syndrome coronavirus 2 among Japanese people unexposed to the virus.

While the COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses a threat to public health as the number of cases and COVID-19-related deaths are increasing worldwide, the incidence of the virus infection is extremely low in Japan compared with many other countries. To explain this uncommon phenomenon, we investigated the prevalence of naturally occurring ("natural") antibodies, focusing on those of the secretory immunoglobulin A (sIgA) form, reactive with SARS-CoV-2 among Japanese people. One hundred and eighty healthy Japanese volunteers of a wide range of age who had been considered to be unexposed to SARS-CoV-2 participated in this study. Saliva samples and blood samples were collected from all of the 180 participants and 139 adults (aged ≥ 20 years) included therein, respectively. The determination of saliva IgA antibodies, mostly comprising sIgA antibodies, as well as serum IgA and immunoglobulin G antibodies, reactive with the receptor binding domain of the SARS-CoV-2 spike-1 subunit proteins was conducted using an enzyme-linked immunosorbent assay. The major findings were that 52.78% (95% confidence interval, 45.21%-60.25%) of the individuals who had not been exposed to SARS-CoV-2 were positive for saliva IgA antibodies with a wide range of levels between 0.002 and 3.272 ng/mL, and that there may be a negative trend in positivity for the antibodies according to age. As we had expected, a frequent occurrence of assumable "natural" sIgA antibodies reactive with SARS-CoV-2 among the studied Japanese participant population was observed.

Epstein-Barr virus-related diffuse large B-cell lymphoma type methotrexate-associated lymphoproliferative disorders presenting in the adrenal gland.

Introduction: Methotrexate-associated lymphoproliferative disorders appear during treatment with methotrexate as an immunosuppressive drug. However, the mechanism and frequency are still unknown, and the treatment is undefined. Case presentation: A 76-year-old woman was admitted to the hospital with back pain, and magnetic resonance imaging showed a tumor in the right adrenal region. She had received methotrexate for rheumatoid arthritis. Enhanced computed tomography showed a tumor of 90 mm in diameter on the dorsal side of the liver abutting to the inferior vena cava. The preoperative diagnosis was a hepatic invasion of right adrenocortical carcinoma and right adrenalectomy was performed. The histopathological diagnosis was diffuse large B-cell lymphoma. The final diagnosis was methotrexate-associated lymphoproliferative disorders. Conclusion: It is important to consider methotrexate-associated lymphoproliferative disorders before surgery when neoplastic lesions are found in patients taking methotrexate.

Glyceraldehyde-derived advanced glycation end-products having pyrrolopyridinium-based crosslinks.

Reducing sugars and reactive aldehydes, such as glyceraldehyde, non-enzymatically react with amino or guanidino groups of proteins to form advanced glycation end-products (AGEs) by the Maillard reaction that involves Schiff base formation followed by Amadori rearrangement. AGEs are found relatively in abundance in the human eye and to accumulate at a higher rate in diseases that impair vision such as cataract, diabetic retinopathy or age-related macular degeneration. We identified two novel AGEs of pyrrolopyridinium lysine dimer derived from glyceraldehyde, PPG1 and PPG2, in the Maillard reaction of N α-acetyl-l-lysine with glyceraldehyde under physiological conditions. Having fluorophores similar to that of vesperlysine A, which was isolated from the human lens, PPGs were found to act as photosensitizers producing singlet oxygen in response to blue light irradiation. Moreover, PPG2 interacts with receptor for AGE (RAGE) in vitro with a higher binding affinity than GLAP, a well-known ligand of the receptor. We also proposed a pathway to form PPGs and discussed how they would be formed in vitro. As glyceraldehyde-derived AGEs have been studied extensively in connection with various hyperglycemia-related diseases, further studies will be required to find PPGs in vivo such as in the lens or other tissues.

A novel crosslinked type of advanced glycation end-product derived from lactaldehyde.

Glycation of amino or guanidino groups of proteins with glucose and glucose-derived reactive aldehydes, such as α-hydroxyaldehydes, leads to accumulation of advanced glycation end-products (AGEs) in the body, resulting in diabetic complications and age-related pathology. Although molecular structures of glycolaldehyde- and glyceraldehyde-derived AGEs have been described in previous studies, little is known about lactaldehyde-derived AGEs of α-hydroxyaldehydes. Here, we report a novel crosslinked type of AGE, named as lactaldehyde-derived lysine dimer (LAK2), which is produced due to non-enzymatic glycation of N α-acetyl-L-lysine with lactaldehyde under physiological conditions. We have identified the molecular structure of LAK2 by extensive mass spectrometry and nuclear magnetic resonance analyses. Furthermore, we propose a reaction pathway to produce LAK2, in which it is formed through an intermediate common with the recently reported lactaldehyde-derived pyridinium-type lysine adduct (LAPL). Since lactaldehyde is known to be produced from L-threonine in a myeloperoxidase (MPO)-mediated reaction at sites of inflammation, LAK2 has the potential to be an oxidative stress marker of MPO-mediated reactions induced in inflammation.

Association between selective IgA deficiency and COVID-19.

The purpose of this study was to propose a hypothesis that there is a potential association between the incidence of selective IgA deficiency in various countries and COVID-19 cases. The number of deaths due to COVID-19 increased in clear proportion to the number of infected patients, and the difference in the number of deaths by country was due to the difference in the number of infected patients. The frequency of selective IgA deficiency has a strong positive correlation with the prevalence of COVID-19 per population. The low infection rate contributed to the low death rate from COVID-19 in Japan, suggesting that the extremely low frequency of selective IgA deficiency may be a contributing factor.

Long-term intake of miso soup decreases nighttime blood pressure in subjects with high-normal blood pressure or stage I hypertension.

The present study aimed to investigate the effects of the combination of Marukome Nenrin miso, which has natriuretic effects, and Marukome MK-34-1 miso, which has potent angiotensin converting enzyme inhibitory effects, on blood pressure (BP) in humans. A total of 40 subjects aged 40-69 years with high-normal BP or stage I hypertension were randomly assigned to two groups: 1) the miso group (32 g 2:1 w/w Nenrin and MK-34-1 with 3.8 g salt/day) or 2) the control soy food group (14.4 g soy food with 0.2 g salt/day). The levels of major nutrients were equal in the miso and control food servings, except for the fiber and Na levels, which were higher in the miso food serving. Daytime and nighttime BP were measured with an automated BP monitor. Compared with the soy food intake, miso intake for 8 weeks did not affect daytime clinical BP but significantly decreased nighttime BP without affecting pulse rate (PR). Moreover, miso shifted the nighttime BP profile to lower levels than those at baseline. Soy food intake did not change the nighttime BP profile after 8 weeks. Miso intake also tended to reduce nighttime BP in a subgroup with stage 1 hypertension compared with the results of the soy food group participants and shifted the nighttime BP profile toward lower levels than those recorded at baseline. Miso intake did not influence lipid or glucose metabolism. In conclusion, this is the first report showing that miso reduces nighttime BP in humans. Miso may do so by shrinking the fluid spaces in the body and/or deactivating the adrenergic nervous system.

Electrical properties of cells from human olfactory epithelium.

OBJECTIVE: The electrical properties of olfactory cells (OCs) are typically examined using animals such as newts, mice, and frogs, with few studies on human OCs. This study investigated the electrical properties of human cells from olfactory epithelium (hCOEs) obtained from subjects of olfactory epithelium showing no clinical symptoms during endoscopic sinus surgery. METHODS: hCOEs were isolated by collagenase treatment for whole-cell patch clamp recording. The identity of the cells was confirmed by immunohistochemistry with an antibody against olfactory maker protein. Under the voltage clamp with the whole-cell recording configuration, the voltage-gated currents of isolated hCOEs were recorded when the membrane potential was depolarized from a holding potential of -100 mV in a stepwise manner between -90 mV and + 40 mV. RESULTS: Only one of 14 hCOE samples expressed a transient inward current at the depolarizing voltage step that was activated by depolarization beyond -40 mV and reached a peak at -30 mV. Delayed and sustained outward currents (444 ± 106 pA at + 40 mV pulse; n = 20) were suppressed by tetraethyl ammonium (n = 3), which is consistent with the properties of newt OCs. CONCLUSIONS: Most hCOEs did not exhibit the transient inward current observed in animal models. These findings provide insight into the physiological basis of the unique aspects of human olfactory signal transduction.

Newly manufactured Marukome MK-34-1 miso with angiotensin-converting enzyme inhibitory activity and its antihypertensive effects in genetic hypertensive rat models.

We newly manufactured miso rich in angiotensin-converting enzyme (ACE) inhibitory activity (Marukome MK-34-1, shinki miso) and investigated its antihypertensive properties in rat models of genetic hypertension. ACE inhibitory activity was tenfold higher in shinki miso than in commercially available Marukome Nenrin miso (nenrin miso). The inhibitory activity of shinki miso was confined to <3 kDa fractions and was detected in several fractions with high polarity by C18 high-performance liquid chromatography. Systolic blood pressure (SBP) increased age-dependently in stroke-prone spontaneously hypertensive rats (SHRSP/Izm) given a 0.6% (w/v) NaCl solution (salt solution group) that matched the salt content of the miso solutions. This SBP increase was attenuated in both the 5% nenrin and 5% shinki miso solution groups compared to the salt solution group. The reduction in SBP was greater in rats fed shinki than in rats fed nenrin miso. Similarly, in a salt-induced hypertension model with Dahl rats, the 5% nenrin miso solution attenuated the rising SBP observed in the salt solution group. Moreover, combining 5% nenrin miso with 5% shinki miso (2:1, v/v) (awase miso group) significantly decreased the SBP per gram salt intake by 8% compared with the nenrin miso treatment. However, there were no differences in urinary Na excretion between the nenrin and awase miso groups. In conclusion, we produced a new miso with potent ACE inhibitory activity that reduced spontaneous and salt-induced hypertension. These results suggest that salt sensitivity is decreased by the addition of shinki miso to nenrin miso.

New photic stimulating system with white light-emitting diodes to elicit electroretinograms from zebrafish larvae.

PURPOSE: The zebrafish is an established animal model commonly used in biological, neuroscience, and genetic research. We have developed a new light stimulating system using white light-emitting diodes (LEDs) to elicit ERGs from zebrafish larvae. The purpose of this study was to record full-field ERGs and to evaluate the inter-trial reliability of the ERGs recorded with our system from zebrafish larvae. METHODS: The stimulating device used white LEDs that were attached to a stereomicroscope, and the location of the recording electrode on the cornea could be monitored while the eye was being stimulated. Full-field scotopic and photopic ERGs were recorded from larvae at the age of 5-7 days post-fertilization (dpf). Intensity-response curves were constructed from the ERGs. Inter-trial reliability of the ERGs recorded by our system was evaluated. RESULTS: This stimulating system could be used for efficient and reliable ERG recordings from 5-7 dpf larvae. The amplitudes, implicit times, and the waveforms of the scotopic and photopic ERGs were similar to those reported in earlier studies. Inter-trial reliability of the amplitudes of the photopic ERG b-waves was excellent with an intra-class correlation coefficient of 0.98. CONCLUSION: We conclude that this new light stimulation system using white LEDs attached to a stereomicroscope will be helpful in recording reliable ERGs from zebrafish larvae.

Monocyte Chemoattractant Protein-1 (MCP-1) as a Potential Therapeutic Target and a Noninvasive Biomarker of Liver Fibrosis Associated With Transient Myeloproliferative Disorder in Down Syndrome.

Liver fibrosis is one of the common complications of transient myeloproliferative disorder (TMD) in Down syndrome (DS), but the exact molecular pathogenesis is largely unknown. We herein report a neonate of DS with liver fibrosis associated with TMD, in which we performed the serial profibrogenic cytokines analyses. We found the active monocyte chemoattractant protein-1 expression in the affected liver tissue and also found that both serum and urinary monocyte chemoattractant protein-1 concentrations are noninvasive biomarkers of liver fibrosis. We also showed a prospective of the future anticytokine therapy with herbal medicine for the liver fibrosis associated with TMD in DS.

Mutation of the key residue for extraribosomal function of ribosomal protein S19 cause increased grooming behaviors in mice.

Ribosomal protein S19 (RP S19) possesses ribosomal function as RP S19 monomer and extraribosomal function as cross-linked RP S19 oligomers which function as a ligand of the complement 5a (C5a) receptor (CD88). We have generated a Gln137Glu-RP S19 knock-in (KI) mouse, which is shown to possess the weakened extraribosomal function of RP S19. Because whether the extraribosomal function of RP S19 has a role in brain function had been unclear, we performed behavioral analysis on these mice and demonstrated that KI mice displayed an increased grooming behavior during open-field test and elevated plus maze test and an enhanced freezing behavior in contextual fear conditioning test. These results suggest an involvement of RP S19 oligomers in some anxiety-like behavior, especially grooming behavior.

Potential of a 70 kDa IL-10-like factor in synovial fluid from rheumatoid arthritis patients to augment superoxide generation by human neutrophils.

AIM: To elucidate the role of polymorphonuclear leukocytes (PMNs) in joint destruction during the inflammatory process in rheumatoid arthritis (RA) as related to superoxide generation. METHODS: Superoxide generation by human peripheral PMNs was measured by using a water-soluble formazan dye, 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2, 4-disulfophenyl)-2H-tetrazolium, monosodium salt, under PMN stimulation with N-formylmethionyl-leucyl-phenylalanine (fMLP) and cytochalasin B. Factors in synovial fluids (SF) from RA patients that may augment PMN superoxide generation were characterized via high-performance liquid chromatography and isoelectric focusing. RESULTS: The formazan dye allowed measurement of superoxide generated in the xanthine-xanthine oxidase system and by PMNs stimulated by cytochalasin B and fMLP in the presence of the intermediate electron transporter phenazine methosulfate. By using chromatography and electrophoresis, an RA-SF protein with an apparent molecular size of 70 kDa and an isoelectric point of 8.3 was isolated and was demonstrated to increase superoxide generation by PMNs. The factor was heat-labile and susceptible to protease treatment. This enhancing activity of the factor was absorbed by human PMNs and was somewhat immunoadsorbed with a specific monoclonal antibody against interleukin (IL)-10. CONCLUSION: The 70-kDa protein factor in RA-SF increased superoxide generation by human PMNs, which suggests the possibility of its being related to IL-10. This factor may have a pathological role in RA joint destruction caused by PMNs and coinciding with rheumatoid inflammation, which suggests that PMNs, via superoxide generation, play an important role in RA joint destruction. IL-10 therefore likely has biological activity toward PMNs during synovial inflammatory chain reactions in RA.

Vascular effects and safety of supplementation with shark liver oil in middle-aged and elderly males.

Shark liver oil (SLO) has long been used as a traditional health food, with a particular benefit for vascular health, in Japan. The aim of this study was to assess the effect of dietary supplementation with SLO on arterial stiffness and peripheral microvascular function in otherwise healthy middle-aged and older males with slightly increased arterial stiffness. A randomized, double-blind, placebo-controlled, parallel study design was used to assign 41 healthy males with a mean age of 59.0±4.0 years (range, 45-69 years) to either SLO (n=21) or placebo (n=20) treatment for eight weeks. The effects on arterial stiffness and peripheral microvascular function were assessed by the cardio-ankle vascular index (CAVI) and by measurement of hand blood flow to cutaneous tissues using a laser Doppler perfusion imaging (LDPI) technique, respectively. Although the magnitude of the changes in the CAVI value during the eight-week intervention for the SLO group did not significantly differ from that for the placebo group, the changes in the CAVI value for the former group were significantly associated (r=0.575, P<0.01) with age. It was also found that the LDPI values at week 8 were significantly lowered (P<0.05) compared with the baseline values in the placebo group, while no change was observed in the SLO group, resulting in a significant difference in the changes between the two groups (P=0.002). Neither SLO supplementation-related adverse side-effects nor any abnormal changes in routine laboratory tests, including lipid profiles and anthropometric and haemodynamic parameters, were observed throughout the intervention. SLO may have the potential to safely improve vascular health in middle-aged and elderly males.

Improvement of liver function by the administration of oyster extract as a dietary supplement to habitual alcohol drinkers: A pilot study.

Alcoholic liver disease (ALD) is characterized by elevated serum γ-glutamyltransferase (GGT) activity with hepatic steatosis, hepatitis or occasionally fibrosis that may progress to cirrhosis. The potential therapeutic role of oyster extract (OE) or OE-containing dietary supplements (OE supplement) in ALD has seldom been evaluated. In the present study, 84 adults who had an alcohol-drinking habit and marginally high serum GGT levels were enrolled in a randomized, double-blind, placebo-controlled feeding trial to study the effect on alcohol-impaired liver function as reflected by an increased serum level of GGT, as well as the safety, of an OE supplement. The subjects were randomized to receive either an OE supplement (OE group) or placebo (placebo group). There were 42 subjects (31 males and 11 females) in each group, and all the enrolled subjects entered the study. Four individuals (5%) dropped out for reasons unassociated with the study and 6 other subjects were excluded from the efficacy analysis because they did not maintain the required frequency of alcohol intake. As a result, 38 subjects in the placebo group and 36 in the OE group underwent efficacy assessment. Assays of GGT and other liver enzymes were performed at baseline (week 0) and at weeks 4, 8 and 12 of the intervention period. The mean serum levels of GGT in the placebo group gradually increased, while those in the OE group tended to decrease, although no significant within-group differences were observed for either group. A significant between-group difference in the change of mean GGT from baseline was, however, found at week 12 (P=0.049). No OE supplement-associated untoward side-effects nor any abnormal changes in routine laboratory tests and anthropometric parameters were observed throughout the 12-week intervention. An OE supplement shows promise in reducing risk factors associated with ALD in adults with an alcohol intake habit.

Predicting early intrahepatic recurrence after curative resection of colorectal liver metastases with molecular markers.

BACKGROUND: The aim of this case-control study was to identify clinicopathological factors and test three relevant biomarkers for their ability to predict early intrahepatic recurrence after curative liver resection for colorectal liver metastases (CLM). METHODS: Of the 184 patients with CLM undergoing hepatectomy between January 2007 and December 2009, thirty patients had intrahepatic disease recurrence within 6 months. The control group was randomly selected from a cohort of patients between April 1997 and December 2005 who have survived without disease recurrence after CLM resection for over 5 years. Both groups were matched for size of metastasis greater than 5.0 cm, the presence of multiple metastases, and synchronous versus metachronous CLM. The final study population consisted of 60 patients with CLM undergoing R0 hepatectomy, 30 of whom had early intrahepatic-only recurrences (study group) and 30 patients without recurrence for more than 5 years (control group). Both groups were analyzed and compared for the presence of clinical factors and expression levels of CD133, survivin, and Bcl-2 within tumor tissue. RESULTS: Characteristics of patients were similar between the two groups except primary tumor location and administration of postoperative chemotherapy. Expression level of CD133 and survivin were significantly increased in tumors of patients with recurrence compared to patients without recurrence. On multivariate analysis high tumor expression levels of CD133 (odds ratio [OR] 14.7, confidence interval [CI] 1.8-121.3, p = 0.012) and survivin (OR 9.5, CI 2.1-44.3, p = 0.004) and postoperative chemotherapy (OR 4.8, CI 1.01-22.9, p = 0.049) were independent factors associated with early intrahepatic recurrence. CONCLUSIONS: Tumor expression levels of CD133 and survivin may be a useful predictor of early intrahepatic recurrence after hepatectomy for CLM. Administration of postoperative chemotherapy may prevent early intrahepatic recurrence.

Clinical impact of gastroenterologist-administered propofol during esophagogastroduodenoscopy: a randomized comparison at a single medical clinic.

BACKGROUND: Although midazolam is widely used during endoscopic procedures by endoscopists, propofol has been recently favored for its rapid action and metabolism. The aim of this study is to compare the clinical advantages between propofol and midazolam use during screening esophagogastroduodenoscopy (EGD) for gastric cancer and post-procedure management at a medical clinic. METHODS: One hundred six healthy patients aged 20-69 years requesting sedation for screening EGD from October 2012 to May 2013 at a single clinic in Japan were randomly assigned to propofol (n = 54) or midazolam (n = 52). Medications were given by bolus injection, and the dose was adjusted by body weight. Sedation level and tolerability during EGD and recovery time were assessed. Sedation level and tolerability were evaluated by American Society of Anesthesiologists responsiveness levels and four levels of the gag reflex, respectively. For safety purposes, endoscopists and nurses were trained in administering propofol and an anesthesiologist was on call at all times. RESULTS: No statistically significant differences were found between the two groups in sedation level and patient tolerability. Full recovery time in the propofol group (4.7 min) was significantly shorter than that in the midazolam group (24 min, P < 0.01). CONCLUSIONS: Regarding post-procedure management of patients in a medical clinic, propofol use might not necessitate a recovery room and excessive assessment tasks because of rapid recovery time without any prolonged reaction, which causes patient compliance. ( CLINICAL TRIAL REGISTRATION NUMBER: UMIN000009142.).