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We report three cases of Waterhouse-Friderichsen syndrome (WFS) that were confirmed during forensic autopsies. Case 1 involved a man in his 50s post-splenectomy. Bacteriological examination revealed Streptococcus pneumoniae (S. pneumonia). The patient was considered to have died of asphyxiation after aspirating vomit. Case 2 involved a man in his 40s. Bacteriological examination again revealed S. pneumoniae. Histopathological examination showed hypoplasia of the spleen. This patient was considered to have died of multiple-organ failure due to sepsis, disseminated intravascular coagulation, and WFS. Case 3 involved a post-splenectomy woman in her 60s with a history of systemic lupus erythematosus. Bacteriological examination revealed Streptococcus oralis. This patient was considered to have died of multiple-organ failure due to sepsis, disseminated intravascular coagulation, and WFS. These three cases were included among forensic autopsies conducted in the last 5 years. WFS has been considered a rare disease, but may be more frequent than previously assumed. If a mildly ill patient displays a sudden change in status and dies within a short period of time, we consider it necessary to perform not only bacteriological examinations, but also histopathological examination of the spleen during autopsy.

Pathway-selective gene delivery would be critical for future gene therapy against neuropsychiatric disorders, traumatic neuronal injuries, or neurodegenerative diseases, because the impaired functions depend on neural circuits affected by the insults. Pathway-selective gene delivery can be achieved by double viral vector techniques, which combine an injection of a retrograde transport viral vector into the projection area of the target neurons and that of an anterograde viral vector into their somas. In this study, we tested the efficiency of gene delivery with different combinations of viral vectors to the pathway extending from the ventral tegmental area (VTA) to the cortical motor regions in rats, considered to be critical in the promotion of motor recovery from neural injuries. It was found that retrograde recombinant adeno-associated virus 2-retro (rAAV2reto) combined with anterograde AAVDJ (type2/type4/type5/type8/type9/avian/bovine/caprine chimera) exhibited the highest transduction efficiency in the short term (3-6 weeks) but high toxicity in the long term (3 months). In contrast, the same rAAV2reto combined with anterograde AAV5 displayed moderate transduction efficiency in the short term but low toxicity in the long term. These data suggest that the combination of anterograde AAV5 and retrograde rAAV2retro is suitable for safe and efficient gene delivery to the VTA-cortical pathway.

To understand the connectome of the axonal arborizations of dopaminergic midbrain neurons, we investigated the anterograde spread of highly sensitive viral tracers injected into the ventral tegmental area (VTA) and adjacent areas in 3 macaques. In 2 monkeys, injections were centered on the lateral VTA with some spread into the substantia nigra, while in one animal the injection targeted the medial VTA with partial spread into the ventro-medial thalamus. Double-labeling with antibodies against transduced fluorescent proteins (FPs) and tyrosine hydroxylase indicated that substantial portions of transduced midbrain neurons were dopaminergic. Interestingly, cortical terminals were found either homogeneously in molecular layer I, or more heterogeneously, sometimes forming patches, in the deeper laminae II-VI. In the animals with injections in lateral VTA, terminals were most dense in somatomotor cortex and the striatum. In contrast, when the medial VTA was transduced, dense terminals were found in dorsal prefrontal and temporal cortices, while projections to striatum were sparse. In all monkeys, orbitofrontal and occipito-parietal cortex received strong and weak innervation, respectively. Thus, the dopaminergic ventral midbrain sends heterogeneous projections throughout the brain. Furthermore, our results suggest the existence of subgroups in meso-dopaminergic neurons depending on their location in the primate ventral midbrain., (© The Author(s) 2021. Published by Oxford University Press.)

Aims/introduction: The objective of the present study was to clarify the association of the type and number of first-degree family history of diabetes (FHD) with the clinical characteristics, especially with residual β-cell function, in type 2 diabetes patients., Materials and Methods: A total of 1,131 type 2 diabetes patients were recruited and divided into four groups according to FHD information as follows: (i) patients without FHD (FHD-); (ii) those with at least one sibling who had diabetes without parental diabetes (FHD+); (iii) those with one parent (FHD++); or (iv) those with both parents (FHD+++) who had diabetes with or without a sibling with diabetes., Results: The percentages of the FHD-, FHD+, FHD++ and FHD+++ groups were 49.4%, 13.4%, 34.0% and 3.2%, respectively. Patients in the FHD++ and FHD+++ groups were significantly younger at the time of diabetes diagnosis (P < 0.001) than those in the FHD- and FHD+ groups, even after adjusting for confounding factors. In addition, the levels of insulin secretion were significantly lower in the patients in the FHD+, FHD++ and FHD+++ groups than those in the FHD- group (P < 0.05) after adjusting for confounding factors, and the patients in the FHD+++ group presented with the lowest levels of insulin secretion among the four groups., Conclusions: Our results showed that in type 2 diabetes patients, the degree of the associations between FHD and clinical characteristics differs according to the number and the type of FHD. In particular, FHD in both parents is most strongly associated with impaired residual β-cell function., (© 2019 The Authors. Journal of Diabetes Investigation published by Asian Association for the Study of Diabetes (AASD) and John Wiley & Sons Australia, Ltd.)

Aim: Among the three adiponectin isoforms, a lower ratio of high molecular weight (HMW) adiponectin to total adiponectin (TA) is well known to cause insulin resistance and type 2 diabetes (T2D). However, how the levels of other adiponectin isoforms, such as the middle molecular weight (MMW) and low molecular weight (LMW) isoforms, and their relative ratio to TA change in T2D subjects has not been determined. Therefore, we investigated the association of these adiponectin-related parameters with T2D., Methods: We examined the associations between adiponectin-related parameters and diabetes in a group of 394 T2D subjects and 374 controls (1st group) randomly selected from among the participants in our previous study. The associations between these parameters and the HOMA-IR in a 2nd group, consisting of the subjects remaining in the 1st group after the exclusion of subjects receiving diabetic medication, were also examined., Result: In the 1st group, after adjusting for confounding factor, the levels of all the adiponectin isoforms and the HMW/TA ratio were significantly lower among the diabetic subjects than among the controls (all P values < 0.01). On the contrary, the LMW/TA ratio was significantly higher among the diabetic subjects (P < 0.01) and was positively associated with T2D (odds ratio = 8.64, P < 0.01). In the 2nd group, the HMW/TA ratio was inversely associated with the HOMA-IR; however, the LMW/TA ratio was positively associated with the HOMA-IR (β for LMW/TA ratio = 0.89, SE = 0.24, P < 0.001), similar to the association with T2D. The MMW/TA ratio was not associated with T2D or the HOMA-IR., Conclusion: The current investigation demonstrated that, unlike the reduction in the levels of all the adiponectin isoforms and the HMW/TA ratio, an increased LMW/TA ratio was associated with T2D through its relation to insulin resistance.

Aim: Several studies have demonstrated that polymorphisms within the fat-mass and obesity-associated gene (FTO) are associated with type 2 diabetes (T2D). However, whether the effects of the FTO locus on T2D susceptibility are independent of fat-mass increases remains controversial. To investigate this issue, we examined the association of FTO variants with T2D and various aspects of BMI history during adult life in a Japanese population., Methods: We genotyped SNPs within FTO (rs1121980 and rs1558902) in 760 Japanese patients with T2D who had reached a lifetime maximum BMI (BMImax) before or at the time of diagnosis and 693 control individuals with information regarding their BMImax., Results: The BMImax showed the strongest association with T2D risk among the BMIs evaluated in this study. In the sex-combined analysis, FTO SNPs were not associated with any of the BMI variables or with T2D, but in sex-stratified analyses, both SNPs were significantly associated with the BMImax and rs1558902 was associated with T2D in men. The association of the SNPs with T2D remained significant after adjustments for the current BMI and age, whereas the T2D association of the SNP was no longer significant after adjustments for BMImax and age., Conclusions: These results suggest that the effects of FTO polymorphisms on T2D susceptibility in Japanese men are mediated through their effect on increasing the BMImax before or at the time of diagnosis., Competing Interests: The authors have declared that no competing interests exist.

Chronic inflammation is a pathophysiology of insulin resistance in metabolic diseases, such as obesity and type 2 diabetes. Adipose tissue macrophages (ATMs) play important roles in this inflammatory process. SIRT1 is implicated in the regulation of glucose metabolism in some metabolic tissues, such as liver or skeletal muscle. This study was performed to investigate whether SIRT1 in macrophages played any roles in the regulation of inflammation and glucose metabolism. Myeloid cell-specific SIRT1-knockout mice were originally generated and analyzed under chow-fed and high-fat-fed conditions. Myeloid cell-specific SIRT1 deletion impaired insulin sensitivity and glucose tolerance assessed by the glucose- or insulin-tolerance test, which was associated with the enhanced expression of inflammation-related genes in epididymal adipose tissue of high-fat-fed mice. Interestingly, the M1 ATMs from the SIRT1-knockout mice showed more hypoxic and inflammatory phenotypes than those from control mice. The expressions of some inflammatory genes, such as Il1b and Nos2 , which were induced by in vitro hypoxia treatment, were further enhanced by SIRT1 deletion along with the increased acetylation of HIF-1α in cultured macrophages. These results suggest that deletion of SIRT1 in myeloid cells impairs glucose metabolism by enhancing the hypoxia and inflammatory responses in ATMs, thereby possibly representing a novel therapeutic target for metabolic diseases, such as type 2 diabetes.

Voltage-gated Na(+) channel β-subunits are multifunctional molecules that modulate Na(+) channel activity and regulate cell adhesion, migration and neurite outgrowth. β-subunits including β4 are known to be highly concentrated in the nodes of Ranvier and axon initial segments in myelinated axons. Here we show diffuse β4 localization in striatal projection fibres using transgenic mice that express fluorescent protein in those fibres. These axons are unmyelinated, forming large, inhibitory fibre bundles. Furthermore, we report β4 dimer expression in the mouse brain, with high levels of β4 dimers in the striatal projection fascicles, suggesting a specific role of β4 in those fibres. Scn4b-deficient mice show a resurgent Na(+) current reduction, decreased repetitive firing frequency in medium spiny neurons and increased failure rates of inhibitory postsynaptic currents evoked with repetitive stimulation, indicating an in vivo channel regulatory role of β4 in the striatum.

Aims/introduction: The objective of the present study was to clarify the validity of β-cell function-related parameters for predicting the insulin requirement of Japanese type 2 diabetic patients., Materials and Methods: In 188 patients with type 2 diabetes who had been admitted to the University of Toyama Hospital (Toyama, Japan) without receiving insulin therapy, we carried out a cross-sectional study examining the relationship between the homeostasis model assessment of β-cell function (HOMA-β) and C-peptide-based indices, and also carried out a retrospective study to examine the utility for predicting insulin requirement of several β -cell function-related indices using a receiver operating characteristic (ROC) curve analysis., Results: The secretory units of islets in transplantation index (SUIT) had the strongest correlation with HOMA-β, followed by the fasting serum C-peptide immunoreactivity index (CPI); the fasting serum C-peptide immunoreactivity itself (F-CPR) had the least correlation. The CPI, HOMA-β and SUIT were significantly lower in the insulin-requiring group than in the non-insulin-requiring group, even after adjustments for confounding factors (P < 0.01). The areas under the ROC curve for insulin requirement were 0.622, 0.774, 0.808, and 0.759 for F-CPR, CPI, SUIT, and HOMA-β, respectively. The cut-off values of SUIT, CPI, and HOMA-β for an over 80% specificity for the prediction of insulin therapy were 23.5, 1.00, and 14.9, respectively., Conclusions: The present study shows that SUIT is the best predictor of insulin requirement among these β-cell function-related markers.

The subthalamic nucleus (STN) of the basal ganglia plays a key role in motor control, and STN efferents are known to mainly target the external segment of the globus pallidus (GPe), entopeduncular nucleus (Ep), and substantia nigra (SN) with some axon collaterals to the other regions. However, it remains to be clarified how each STN neuron projects axon fibers and collaterals to those target nuclei of the STN. Here we visualized the whole axonal arborization of single STN neurons in the rat brain by using a viral vector expressing membrane-targeted green fluorescent protein, and examined the distribution of axon boutons in those target nuclei. The vast majority (8-9) of 10 reconstructed STN neurons projected to the GPe, SN, caudate-putamen (CPu), and Ep, which received, on average ± SD, 457 ± 425, 400 ± 347, 126 ± 143, and 106 ± 100 axon boutons per STN neuron, respectively. Furthermore, the density of axon boutons in the GPe was highest among these nuclei. Although these target nuclei were divided into calbindin-rich and -poor portions, STN projection showed no exclusive preference for those portions. Since STN neurons mainly projected not only to the GPe, SN, and Ep but also to the CPu, the subthalamostriatal projection might serve as a positive feedback path for the striato-GPe-subthalamic disinhibitory pathway, or work as another route of cortical inputs to the striatum through the corticosubthalamostriatal disynaptic excitatory pathway., (Copyright © 2012 Wiley Periodicals, Inc.)

Background and Objective: In asthma, reduced histone deacetylase activity and enhanced histone acetyltransferase activity in the lungs have been reported. However, the precise function of Sirtuin 1 (Sirt1), a class III histone deacetylase, and the effect of the Sirt1 activator SRT1720 on allergic inflammation have not been fully elucidated., Methods: The effect of SRT1720, a synthetic activator of Sirt1, in an ovalbumin (OVA)-induced asthma mouse model was investigated. The effect of SRT1720 and resveratrol on OVA stimulation in splenocytes from OVA-sensitized and challenged mice was also examined., Results: In OVA-sensitized and challenged mice (OVA mice) compared with saline-sensitized and challenged mice (control mice), Sirt1 messenger RNA expression in the lungs was decreased (P = 0.02), while cellular infiltration, airway eosinophilia and bronchoalveolar lavage (BAL) fluid levels of interleukin (IL)-4, IL-5 and IL-13 were increased (P < 0.01). In OVA mice, SRT1720 treatment decreased total and eosinophil cell counts and IL-5 and IL-13 levels in the BAL fluid compared with the vehicle treatment (P < 0.05). In OVA mice, SRT1720 treatment also decreased inflammatory cell lung infiltrates histologically (P = 0.002). Both SRT1720 and resveratrol suppressed OVA-induced cell proliferation and IL-6 (P < 0.05) and tumour necrosis factor-α (TNF-α) (P < 0.05) production in splenocytes (P < 0.01)., Conclusions: The Sirt1 activator SRT1720 suppressed inflammatory cell infiltration and cytokine production in an OVA-induced mouse model of asthma. SRT1720 and resveratrol suppressed OVA-induced splenocyte proliferation and TNF-α and IL-6 production. Sirt1 activators might have beneficial effects in asthmatics by suppressing inflammation., (© 2012 The Authors. Respirology © 2012 Asian Pacific Society of Respirology.)

Background: The developmental mechanisms of breast neuroendocrine carcinoma (B-NEC) have not been sufficiently analysed and are not well understood., Aims: To investigate NE cells in the background tissues surrounding B-NECs., Methods: Three cases (four breasts) having many NE cells in the background tissues of multifocal B-NECs were identified at the University of Yamanashi Hospital and St Luke's International Hospital, Japan. These patients were, respectively, 28-, 31- and 38-year-old women with no familial history of NE tumour. The totally-resected breasts were serially studied by immunohistochemistry for specific NE markers (chromogranin A/synaptophysin) and the morphologies and/or localisation of NE cells were investigated., Results: Immunohistochemical examination showed extensively-distributed NE cells in the background mammary ducts/lobules of the NECs in all breasts. These NE cells were classifiable into three emerging patterns: isolated/scattered, clustered and circumferential. Their distributions were intermingled and were not clearly related to B-NEC foci. NE cells were morphologically polygonal, oval or columnar with sometimes eosinophilic and/or fine-granular cytoplasm and round-to-ovoid nuclei lacking atypia. Some cells were located between epithelial and myoepithelial cells. Apical snouts were occasionally observed in NE cells forming luminal structures., Conclusions: Benign-appearing NE cells in the parenchyma of a breast with NEC could be regarded as hyperplastic from their emerging patterns and distribution; this NE cell hyperplasia may be associated with the histogenesis of B-NEC as a precancerous condition. These observations might raise questions about the treatment for B-NEC.

Leukocyte cell-derived chemotaxin 2 (LECT2) facilitates neuritic extension in cultured hippocampal neurons during initial development. However, the functions of LECT2 in neuritic extension are poorly understood. To elucidate these functions, we examined microtubular morphology and levels of katanin-P60, a microtubule-severing enzyme, in cultured hippocampal neurons from wild-type mice and LECT2 knockout (KO) mice. Microtubules in LECT2-KO mice exhibited fragmentation and were shorter than those of wild-type mice. Furthermore, the expression of katanin-P60 in LECT2-KO mice was significantly elevated when compared with wild-type mice at 1 day in vitro (DIV1) and 4. Our results suggest that LECT2 regulates neuritic extension through microtubular morphallaxis through the control of katanin-P60 levels.

Enkephalin (ENK) has been implicated in nociceptive transmission in the spinal cord while its functional role is not clear because of difficulties in ideally visualizing ENKergic neurons. We thus developed preproenkephalin-green fluorescent protein transgenic mice to better identify ENKergic neurons. Real-time reverse transcriptase-polymerase chain reaction (RT-PCR) together with immunohistochemistry and in situ hybridization were first employed to confirm the successful transgenic manipulation and its application in showing spinal ENKergic neurons. The proportions of ENKergic neurons in the spinal cord laminae I, II, III and IV-VI among dorsal horn neurons were 15.8 +/- 3.1%, 39.5 +/- 3.3%, 11.8 +/- 1.9% and 10.7 +/- 2.1%, respectively. Double labeling with other molecules was then performed to further clarify the neurochemical properties of spinal ENKergic neurons. GABA was found to exist in 42.9 +/- 2.8% of ENKergic neurons that were mainly located in lamina I-III. The proportions of parvalbumin-, calretinin-, calbindin- and neuronal nitric oxide synthase-positive cells among the ENKergic neurons were 5.2 +/- 0.7%, 42.6 +/- 2.3%, 25.8 +/- 2.2% and 11.1 +/- 1.6%, respectively. Compared with previously findings obtained with ENK antibody labeling, this line of newly generated mice can be a reliable tool for the study of specific spinal ENKergic neuronal population.

Leukocyte cell-derived chemotaxin 2 (LECT2) was first isolated as a chemotactic factor from phytohemagglutinin-activated human T-cell leukemia SKW-3 cells. LECT2 is expressed in various tissues, including in the brain, stomach and liver, but the functions of LECT2 in the brain remains unclear. To elucidate these functions, we investigated the influence of a deficiency of LECT2 on the morphology of cultured hippocampal neurons during neuronal development, and examined the expression of neurotrophins (NGF, BDNF, and NT-3) and their receptors (TrkA, TrkB, TrkC, and p75NTR) in these neurons. The extension of axons and dendrites in neurons from LECT2-knockout (LECT2-KO) mice was shorter than that in neurons from wild-type mice during culture and significantly less than that in wild-type mice after 4 days in culture. Moreover, neurons from LECT2-KO mice showed different expression of NGF, BDNF and NT-3 during culture compared to wild-type mice. Our results show that LECT2 regulates the extension of axons and dendrites and the expressions of NGF, BDNF and NT-3 during neuronal development., (Copyright 2009 Elsevier B.V. All rights reserved.)

Whether or not the striosome compartment of the neostriatum contained preproenkephalin (PPE)-expressing neurons remained unresolved. To address this question by developing a sensitive detection method, we generated transgenic mice expressing enhanced green fluorescent protein (GFP) under the specific transcriptional control of the PPE gene. Eight transgenic lines were established, and three of them showed GFP expression which was distributed in agreement with the reported localization of PPE mRNA in the central nervous system. Furthermore, in the matrix compartment of the neostriatum of the three lines, intense GFP immunoreactivity was densely distributed in the neuronal cell bodies and neuropil, and matrix neurons displayed > 94% co-localization for GFP and PPE immunoreactivities. In sharp contrast, GFP immunoreactivity was very weak in the striosome compartment, which was characterized by intense immunoreactivity for mu-opioid receptors (MOR). Although neostriatal neurons were divided into GFP-immunopositive and -negative groups in both the striosome and matrix compartments, GFP immunoreactivity of cell bodies was much weaker (~1/5) in GFP-positive striosomal neurons than in GFP-positive matrix neurons. A similar reciprocal organization of PPE and MOR expression was also suggested in the ventral striatum, because GFP immunoreactivity was weaker in intensely MOR-immunopositive regions than in the surrounding MOR-negative regions. As PPE-derived peptides are endogenous ligands for MOR in the neostriatum and few axon collaterals of matrix neurons enter the striosome compartment, the present results raised the question of the target of those peptides produced abundantly by matrix neurons.

Enhanced green fluorescent protein (GFP) irreversibly loses not only fluorescence but also antigenicity recognized with conventional anti-GFP antibodies by heat denaturation. This hinders combinatory applications of the GFP immunodetection technique with heat-requiring procedures, such as in situ hybridization histochemistry, antigen retrieval, and Western blot. Here we produced new rabbit and guinea pig antibodies against heat-denatured GFP. The polyclonal antibodies affinity-purified with the antigen column detected a single band corresponding to the molecular size of GFP in Western blot analysis, with mouse brain expressing GFP from the GAD67 locus. By immunofluorescence labeling, the new antibodies detected GFP molecules in heat (> or = 70 degrees C)-treated sections but not in untreated sections of the mouse brain. When the sections were incubated at > or = 37 degrees C with in situ hybridization buffer containing 50% formamide, a denaturing reagent, the sections lost immunoreactivity with the conventional anti-GFP antibodies but acquired immunoreactivity with the new antibodies to heat-denatured GFP. Finally, GFP immunofluorescence was successfully visualized with the new antibodies in sections of the GFP-expressing mice labeled by fluorescence in situ hybridization histochemistry against GAD67 mRNA. Thus, the antibodies produced in this study may provide an opportunity to combine GFP immunodetection with procedures requiring heat treatment. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.

Expression of vesicular glutamate transporters (VGLUTs: VGLUT1, VGLUT2 and VGLUT3) in muscle spindle afferents was examined in rats. VGLUT1 immunoreactivity was detected in the sensory endings on the equatorial and juxta-equatarial regions of intrafusal fibers as well as in many axon terminals within lamina IX of the spinal cord. VGLUT1 might be expressed not only in the central axon terminals but also in the peripheral sensory endings of muscle-spindle afferents., (Copyright 2004 Elsevier B.V.)