The brain-specific kinase LMTK3 regulates neuronal excitability by decreasing KCC2-dependent neuronal Cl- extrusion.

LMTK3 is a brain-specific transmembrane serine/threonine protein kinase that acts as a scaffold for protein phosphatase-1 (PP1). Although LMKT3 has been identified as a risk factor for autism and epilepsy, its physiological significance is unknown. Here, we demonstrate that LMTK3 copurifies and binds to KCC2, a neuron-specific K+/Cl- transporter. KCC2 activity is essential for Cl--mediated hyperpolarizing GABAAR receptor currents, the unitary events that underpin fast synaptic inhibition. LMTK3 acts to promote the association of KCC2 with PP1 to promote the dephosphorylation of S940 within its C-terminal cytoplasmic domain, a process the diminishes KCC2 activity. Accordingly, acute inhibition of LMTK3 increases KCC2 activity dependent upon S940 and increases neuronal Cl- extrusion. Consistent with this, LMTK3 inhibition reduced intrinsic neuronal excitability and the severity of seizure-like events in vitro. Thus, LMTK3 may have profound effects on neuronal excitability as an endogenous modulator of KCC2 activity.

Memory CD8 T cells are vulnerable to chronic IFN-γ signals but not to CD4 T cell deficiency in MHCII-deficient mice.

The mechanisms by which the number of memory CD8 T cells is stably maintained remains incompletely understood. It has been postulated that maintaining them requires help from CD4 T cells, because adoptively transferred memory CD8 T cells persist poorly in MHC class II (MHCII)-deficient mice. Here we show that chronic interferon-γ signals, not CD4 T cell-deficiency, are responsible for their attrition in MHCII-deficient environments. Excess IFN-γ is produced primarily by endogenous colonic CD8 T cells in MHCII-deficient mice. IFN-γ neutralization restores the number of memory CD8 T cells in MHCII-deficient mice, whereas repeated IFN-γ administration or transduction of a gain-of-function STAT1 mutant reduces their number in wild-type mice. CD127high memory cells proliferate actively in response to IFN-γ signals, but are more susceptible to attrition than CD127low terminally differentiated effector memory cells. Furthermore, single-cell RNA-sequencing of memory CD8 T cells reveals proliferating cells that resemble short-lived, terminal effector cells and documents global downregulation of gene signatures of long-lived memory cells in MHCII-deficient environments. We propose that chronic IFN-γ signals deplete memory CD8 T cells by compromising their long-term survival and by diverting self-renewing CD127high cells toward terminal differentiation.

Mass spectrometry-based proteomic analysis of proteins adsorbed by hexadecyl-immobilized cellulose bead column for the treatment of dialysis-related amyloidosis.

BACKGROUND: Dialysis-related amyloidosis (DRA) is a severe complication in end-stage kidney disease (ESKD) patients undergoing long-term dialysis treatment, characterized by the deposition of ß2-microglobulin-related amyloids (Aß2M amyloid). To inhibit DRA progression, hexadecyl-immobilized cellulose bead (HICB) columns are employed to adsorb circulating ß2-microglobulin (ß2M). However, it is possible that the HICB also adsorbs other molecules involved in amyloidogenesis. METHODS: We enrolled 14 ESKD patients using HICB columns for DRA treatment; proteins were extracted from HICBs following treatment and identified using liquid chromatography-linked mass spectrometry. We measured the removal rate of these proteins and examined the effect of those molecules on Aß2M amyloid fibril formation in vitro. RESULTS: We identified 200 proteins adsorbed by HICBs. Of these, 21 were also detected in the amyloid deposits in the carpal tunnels of patients with DRA. After passing through the HICB column and hemodialyzer, the serum levels of proteins such as ß2M, lysozyme, angiogenin, complement factor D and matrix Gla protein were reduced. These proteins acted in the Aß2M amyloid fibril formation. CONCLUSIONS: HICBs adsorbed diverse proteins in ESKD patients with DRA, including those detected in amyloid lesions. Direct hemoperfusion utilizing HICBs may play a role in acting Aß2M amyloidogenesis by reducing the amyloid-related proteins.

Silencing dentate newborn neurons alters excitatory/inhibitory balance and impairs behavioral inhibition and flexibility.

Adult neurogenesis confers the hippocampus with unparalleled neural plasticity, essential for intricate cognitive functions. The specific influence of sparse newborn neurons (NBNs) in modulating neural activities and subsequently steering behavior, however, remains obscure. Using an engineered NBN-tetanus toxin mouse model (NBN-TeTX), we noninvasively silenced NBNs, elucidating their crucial role in impulse inhibition and cognitive flexibility as evidenced through Morris water maze reversal learning and Go/Nogo task in operant learning. Task-based functional MRI (tb-fMRI) paired with operant learning revealed dorsal hippocampal hyperactivation during the Nogo task in male NBN-TeTX mice, suggesting that hippocampal hyperexcitability might underlie the observed behavioral deficits. Additionally, resting-state fMRI (rs-fMRI) exhibited enhanced functional connectivity between the dorsal and ventral dentate gyrus following NBN silencing. Further investigations into the activities of PV+ interneurons and mossy cells highlighted the indispensability of NBNs in maintaining the hippocampal excitation/inhibition balance. Our findings emphasize that the neural plasticity driven by NBNs extensively modulates the hippocampus, sculpting inhibitory control and cognitive flexibility.

Predicting Intracardiac Thrombus Formation in the Left Atrial Appendage Using Machine Learning and CT Images.

This paper proposes a comprehensive method for estimating thrombus formation factors in the left atrial appendage (LAA). First, using 3D CT (Computer Tomography) image data as input, classification of thrombus presence/absence is learned using 3D ResNet. Besides, 3D Grad-CAM is applied to the prediction results to visualize regions of interest in thrombus formation. Second, features are extracted based on the visualization of regions of interest. Using the extracted features and numerical data obtained from the hospital as input, a regression analysis is performed to predict the presence/absence of thrombus using LightGBM. Visualization of regions of interest using 3D ResNet and 3D Grad-CAM shows that the right inferior pulmonary vein and the LAA were particularly correlated with thrombus formation. Estimation of important factors for thrombus formation using LightGBM shows that the LAA ostium area has the greatest influence on thrombus formation.Clinical Relevance-This paper shows the factors that contribute to thrombus formation in the LAA from the viewpoint of three-dimensional structure. In addition, the features considered important in thrombus formation were identified by comparing a variety of features.

Ligneous periodontitis exacerbated by Behçet's disease in a patient with plasminogen deficiency and a stop-gained variant PLG c.1468C > T: a case report.

BACKGROUND: Plasminogen serves as the precursor to plasmin, an essential element in the fibrinolytic process, and is synthesized primarily in the liver. Plasminogen activation occurs through the action of plasminogen activator, converting it into plasmin. This conversion greatly enhances the fibrinolytic system within tissues and blood vessels, facilitating the dissolution of fibrin clots. Consequently, congenital deficiency of plasminogen results in impaired fibrin degradation. Patients with plasminogen deficiency typically exhibit fibrin deposits in various mucosal sites throughout the body, including the oral cavity, eyes, vagina, and digestive organs. Behcet's disease is a chronic recurrent systemic inflammatory disease with four main symptoms: aphthous ulcers of the oral mucosa, vulvar ulcers, skin symptoms, and eye symptoms, and has been reported worldwide. This disease is highly prevalent around the Silk Road from the Mediterranean to East Asia. We report a case of periodontitis in a patient with these two rare diseases that worsened quickly, leading to alveolar bone destruction. Genetic testing revealed a novel variant characterized by a stop-gain mutation, which may be a previously unidentified etiologic gene associated with decreased plasminogen activity. CASE PRESENTATION: This case report depicts a patient diagnosed with ligneous gingivitis during childhood, originating from plasminogen deficiency and progressing to periodontitis. Genetic testing revealed a suspected association with the PLG c.1468C > T (p.Arg490*) stop-gain mutation. The patient's periodontal condition remained stable with brief intervals of supportive periodontal therapy. However, the emergence of Behçet's disease induced acute systemic inflammation, necessitating hospitalization and treatment with steroids. During hospitalization, the dental approach focused on maintaining oral hygiene and alleviating contact-related pain. The patient's overall health improved with inpatient care and the periodontal tissues deteriorated. CONCLUSIONS: Collaborative efforts between medical and dental professionals are paramount in comprehensively evaluating and treating patients with intricate complications from rare diseases. Furthermore, the PLG c.1468C > T (p.Arg490*) stop-gain mutation could contribute to the association between plasminogen deficiency and related conditions.

Lethal complication: Ventricular septal perforation and right ventricular infarction after acute myocardial infarction.

Here, we report a case of ventricular septal perforation complicated with right ventricular infarction after inferior acute myocardial infarction, which was associated with a poor clinical outcome despite the successful surgical treatment.

The CCR4-NOT complex suppresses untimely translational activation of maternal mRNAs.

Control of mRNA poly(A) tails is essential for regulation of mRNA metabolism, specifically translation efficiency and mRNA stability. Gene expression in maturing oocytes relies largely on post-transcriptional regulation, as genes are transcriptionally silent during oocyte maturation. The CCR4-NOT complex is a major mammalian deadenylase, which regulates poly(A) tails of maternal mRNAs; however, the function of the CCR4-NOT complex in translational regulation has not been well understood. Here, we show that this complex suppresses translational activity of maternal mRNAs during oocyte maturation. Oocytes lacking all CCR4-NOT deadenylase activity owing to genetic deletion of its catalytic subunits, Cnot7 and Cnot8, showed a large-scale gene expression change caused by increased translational activity during oocyte maturation. Developmental arrest during meiosis I in these oocytes resulted in sterility of oocyte-specific Cnot7 and Cnot8 knockout female mice. We further showed that recruitment of CCR4-NOT to maternal mRNAs is mediated by the 3'UTR element CPE, which suppresses translational activation of maternal mRNAs. We propose that suppression of untimely translational activation of maternal mRNAs via deadenylation by CCR4-NOT is essential for proper oocyte maturation.

microRNAs slow translating ribosomes to prevent protein misfolding in eukaryotes.

Slower translation rates reduce protein misfolding. Such reductions in speed can be mediated by the presence of non-optimal codons, which allow time for proper folding to occur. Although this phenomenon is conserved from bacteria to humans, it is not known whether there are additional eukaryote-specific mechanisms which act in the same way. MicroRNAs (miRNAs), not present in prokaryotes, target both coding sequences (CDS) and 3' untranslated regions (UTR). Given their low suppressive efficiency, it has been unclear why miRNAs are equally likely to bind to a CDS. Here, we show that miRNAs transiently stall translating ribosomes, preventing protein misfolding with little negative effect on protein abundance. We first analyzed ribosome profiles and miRNA binding sites to examine whether miRNAs stall ribosomes. Furthermore, either global or specific miRNA deficiency accelerated ribosomes and induced aggregation of a misfolding-prone polypeptide reporter. These defects were rescued by slowing ribosomes using non-cleaving shRNAs as miRNA mimics. We finally show that proinsulin misfolding, associated with type II diabetes, was resolved by non-cleaving shRNAs. Our findings provide a eukaryote-specific mechanism of co-translational protein folding and a previously unknown mechanism of action to target protein misfolding diseases.

Microheterogeneity and Individual Differences of Human Urinary N-Glycome under Normal Physiological Conditions.

Urine is considered an outstanding biological fluid for biomarker discovery, reflecting both systemic and urogenital physiology. However, analyzing the N-glycome in urine in detail has been challenging due to the low abundance of glycans attached to glycoproteins compared to free oligosaccharides. Therefore, this study aims to thoroughly analyze urinary N-glycome using LC-MS/MS. The N-glycans were released using hydrazine and labeled with 2-aminopyridine (PA), followed by anion-exchange fractionation before LC-MS/MS analysis. A total of 109 N-glycans were identified and quantified, of which 58 were identified and quantified repeatedly in at least 80% of samples and accounted for approximately 85% of the total urinary glycome signal. Interestingly, a comparison between urine and serum N-glycome revealed that approximately 50% of the urinary glycome could originate from the kidney and urinary tract, where they were exclusively identified in urine, while the remaining 50% were common in both. Additionally, a correlation was found between age/sex and the relative abundances of urinary N-glycome, with more age-related changes observed in women than men. The results of this study provide a reference for human urine N-glycome profiling and structural annotations.

Urinary proteome profiles associated with cognitive decline in community elderly residents-A pilot study.

Non-invasive and simple methods enabling easy identification of individuals at high risk of cognitive decline are needed as preventive measures against dementia. This pilot study aimed to explore protein biomarkers that can predict cognitive decline using urine, which can be collected non-invasively. Study subjects were selected from participants in a cohort study of middle-aged and older community-dwelling adults who underwent cognitive testing using the Mini-Mental State Examination and provided spot urine samples at two time points with an interval of approximately 5 years. Seven participants whose cognitive function declined 4 or more points from baseline (Group D) and 7 sex- and age-matched participants whose cognitive function remained within the normal range during the same period (Group M) were selected. Urinary proteomics using mass spectrometry was performed and discriminant models were created using orthogonal partial least squares-discriminant analysis (OPLS-DA). OPLS-DA yielded two models that significantly discriminated between the two groups at baseline and follow-up. Both models had ORM1, ORM2, and SERPINA3 in common. A further OPLS-DA model using baseline ORM1, ORM2, and SERPINA3 data showed similar predictive performance for data at follow-up as it did for baseline data (sensitivity: 0.85, specificity: 0.85), with the receiver operating characteristic curve analysis yielding an area under the curve of 0.878. This prospective study demonstrated the potential for using urine to identify biomarkers of cognitive decline.

Personalized Preclinical Training in Dental Ergonomics and Endodontics in Undergraduate Dentistry Students (Pilot Study).

The curriculum at the Department of Pathophysiology in the Periodontal Sciences program at Okayama University includes normative preclinical training (NPT) using phantoms. NPT is given to the whole class of 5 th year students divided in groups of 8 students/instructor. In 2019, an innovative personalized preclinical training (PPT) pilot study was implemented for this group of students whereby two students, each with their own dental unit, were coached by one instructor. The main topics covered were dental ergonomics and endodontics. We aimed to evaluate the effectiveness of PPT in dental ergonomics and endodontics toward increasing the knowledge and future clinical skills of students who had already undergone NPT. A test on endodontics was taken before and after PPT. A questionnaire was completed to assess their perception of improvement regarding the above-mentioned topics. Test scores and questionnaire results both showed that the students' level of knowledge and awareness of future clinical skills was significantly higher after PPT. This pilot study demonstrated that PPT increased the students' knowledge and future clinical skills. As preclinical training forms the foundation for clinical practice, investment in future research regarding this personalized approach is likely to enhance students' understanding and clinical performance.

The Fungal Metabolite (+)-Terrein Abrogates Inflammatory Bone Resorption via the Suppression of TNF-α Production in a Ligature-Induced Periodontitis Mouse Model.

Current periodontal treatment focuses on the mechanical removal of the source of infection, such as bacteria and their products, and there is no approach to control the host inflammatory response that leads to tissue destruction. In order to control periodontal inflammation, we have previously reported the optimization of (+)-terrein synthesis methods and the inhibitory effect of (+)-terrein on osteoclast differentiation in vitro. However, the pharmacological effect of (+)-terrein in vivo in the periodontitis model is still unknown. In this study, we investigated the effect of synthetic (+)-terrein on inflammatory bone resorption using a ligature-induced periodontitis mouse model. Synthetic (+)-terrein (30 mg/kg) was administered intraperitoneally twice a week to the mouse periodontitis model. The control group was treated with phosphate buffer. One to two weeks after the induction of periodontitis, the periodontal tissues were harvested for radiological evaluation (micro-CT), histological evaluation (HE staining and TRAP staining), and the evaluation of inflammatory cytokine production in the periodontal tissues and serum (quantitative reverse-transcription PCR, ELISA). The synthetic (+)-terrein-treated group suppressed alveolar bone resorption and the number of osteoclasts in the periodontal tissues compared to the control group (p < 0.05). In addition, synthetic (+)-terrein significantly suppressed both mRNA expression of TNF-α in the periodontal tissues and the serum concentration of TNF-α (both p < 0.05). In conclusion, we have demonstrated that synthetic (+)-terrein abrogates alveolar bone resorption via the suppression of TNF-α production and osteoclast differentiation in vivo. Therefore, we could expect potential clinical effects when using (+)-terrein on inflammatory bone resorption, including periodontitis.

Regulation mechanisms of CARMA1-Bcl10-MALT1 complex assembly inferred from the analysis of TRAF6-deficient cells.

The CARMA1-Bcl10-MALT1 (CBM) signalosome is a crucial module of NF-κB activation in B cell receptor (BCR) signaling. Biophysical studies have shown that the E3 ubiquitin ligase TRAF6 cooperatively modifies the CBM signalosome; however, the specific details regarding how TRAF6 is involved in BCR signal-induced CBM formation remain unclear. In this study, we aimed to reveal the influences of TRAF6 on CBM formation and TAK1 and IKK activities using DT40 B cells which lack all the exons of TRAF6. In TRAF6-null cells we found: (i) attenuation of TAK1 activity and abolishment of IKK activity and (ii) sustained binding of CARMA1 to Bcl10. To account for the molecular mechanism causing these dynamics, we performed a mathematical model analysis. The mathematical model analysis showed that the regulation of IKK activation by TRAF6 can reproduce TAK1 and IKK activities in TRAF6 null cells, and that the TRAF6 related signal-dependent inhibitor suppresses CARMA1 binding to Bcl10 in wild-type cells. These results suggest that TRAF6 contributes to the positive regulation of IKK activation via TAK1, alongside the negative signal-dependent regulation of CARMA1 binding to Bcl10.

Age distribution and disease severity of COVID-19 patients continued to change in a time-dependent manner from May 2021 to April 2022 in the regional core hospital in Japan.

The present retrospective study aimed to examine the real-world data regarding time-dependent changes in the age distribution of patients with coronavirus disease 2019 (COVID-19) as well as the severity and infectivity in a regional core hospital in Japan. Patients with COVID-19 who visited the fever outpatient branch in Takagi Hospital during phase I (May 1 to December 31, 2021), and during phase II (January 1 to April 30, 2022) were evaluated. The age distribution of outpatients and the characteristics of inpatients aged > 75 years were compared between phases I and II. The age distribution of outpatients shifted from the older generation in phase I to the younger generation in phase II (p < 0.01). Disease severity might be reduced in a time-dependent manner with a decrease in the hospitalization rate (phase I: 145/368 (39.4%); phase II: 104/1496 (7.0%); p < 0.01) and mortality rate (phase I: 10/368 (2.7%); phase II: 7/1496 (0.5%); p < 0.01). The number of patients increased in phase II (374.0/month) compared to that in phase I (36.8/month). Regarding the older inpatients, the disease severity of COVID-19 and hospitalization days were reduced in phase II compared to those in phase I (p < 0.01, each). In conclusion, the present study suggests a change in the age distribution of patients with COVID-19, a decrease in toxicity, and an increase in infectivity of severe acute respiratory syndrome coronavirus 2 in a time-dependent manner.

Reattachment of Fractured Tooth Fragment by Multidisciplinary Treatment Approach.

Dental anterior fractures are common injuries, especially in those who practice extreme sports. This report describes a 25-year-old Bolivian patient who attended our private dental clinic in La Paz, Bolivia after experiencing an accident during downhill mountain biking. An intraoral examination revealed a fracture line on the buccal side in the middle third of the coronal portion of the right central maxillary incisor which extended towards the proximal and lingual sides. Multidisciplinary treatment, including crown lengthening, osteotomy, root canal treatment, fiberglass post insertion, and reattachment of the fracture segment was performed. A follow-up examination at 10 months later revealed that the tooth was completely reestablished both functionally and esthetically and that there was no periapical pathosis or discomfort. This outcome suggests that if a patient seeks a dental consultation soon after a complex crown-root fracture has occurred, and if the broken tooth segment is available, then reattachment offers an economical and simple treatment option which will allow immediate restoration of functionality and esthetic standards. Continued follow-up should form part of such a treatment plan to allow long-term pulp vitality and periodontal health status to be monitored.

Establishing protein expression profiles involved in tooth development using a proteomic approach.

Various growth and transcription factors are involved in tooth development and developmental abnormalities; however, the protein dynamics do not always match the mRNA expression level. Using a proteomic approach, this study comprehensively analyzed protein expression in epithelial and mesenchymal tissues of the tooth germ during development. First molar tooth germs from embryonic day 14 and 16 Crlj:CD1 (ICR) mouse embryos were collected and separated into epithelial and mesenchymal tissues by laser microdissection. Mass spectrometry of the resulting proteins was carried out, and three types of highly expressed proteins [ATP synthase subunit beta (ATP5B), receptor of activated protein C kinase 1 (RACK1), and calreticulin (CALR)] were selected for immunohistochemical analysis. The expression profiles of these proteins were subsequently evaluated during all stages of amelogenesis using the continuously growing incisors of 3-week-old male ICR mice. Interestingly, these three proteins were specifically expressed depending on the stage of amelogenesis. RACK1 was highly expressed in dental epithelial and mesenchymal tissues during the proliferation and differentiation stages of odontogenesis, except for the pigmentation stage, whereas ATP5B and CALR immunoreactivity was weak in the enamel organ during the early stages, but became intense during the maturation and pigmentation stages, although the timing of the increased protein expression was different between the two. Overall, RACK1 plays an important role in maintaining the cell proliferation and differentiation in the apical end of incisors. In contrast, ATP5B and CALR are involved in the transport of minerals and the removal of organic materials as well as matrix deposition for CALR.

Autophagy as a potential mechanism underlying the biological effect of 1,25-Dihydroxyvitamin D3 on periodontitis: a narrative review.

The major active form of vitamin D, 1,25-dihydroxyvitamin D3 (1,25D3), is known for its wide bioactivity in periodontal tissues. Although the exact mechanisms underlying its protective action against periodontitis remain unclear, recent studies have shown that 1,25D3 regulates autophagy. Autophagy is vital for intracellular pathogen invasion control, inflammation regulation, and bone metabolic balance in periodontal tissue homeostasis, and its regulation could be an interesting pathway for future periodontal studies. Since vitamin D deficiency is a worldwide health problem, its role as a potential regulator of autophagy provides new insights into periodontal diseases. Based on this premise, this narrative literature review aimed to investigate the possible connection between 1,25D3 and autophagy in periodontitis. A comprehensive literature search was conducted on PubMed using the following keywords (e.g., vitamin D, autophagy, periodontitis, pathogens, epithelial cells, immunity, inflammation, and bone loss). In this review, the latest studies on the protective action of 1,25D3 against periodontitis and the regulation of autophagy by 1,25D3 are summarized, and the potential role of 1,25D3-activated autophagy in the pathogenesis of periodontitis is analyzed. 1,25D3 can exert a protective effect against periodontitis through different signaling pathways in the pathogenesis of periodontitis, and at least part of this regulatory effect is achieved through the activation of the autophagic response. This review will help clarify the relationship between 1,25D3 and autophagy in the homeostasis of periodontal tissues and provide perspectives for researchers to optimize prevention and treatment strategies in the future.

Primary diffuse large B cell lymphoma of the prostate in a patient with HIV infection.

Introduction: Primary prostate lymphomas are very rare; however, the incidence of malignant lymphoma is high among HIV-infected patients. Herein, we report a case of primary diffuse large B-cell lymphoma (DLBCL) of the prostate in an HIV-infected patient. Case presentation: A 47-year-old man presented with miction pain and back pain. Abdominal CT revealed a huge prostate mass extending to the left retroperitoneum. Serum sIL-2R level was abnormally high (2896 U/mL), whereas PSA level was normal. HIV antigen and antibody tests were positive. The patient was diagnosed with DLBCL after a prostate biopsy. Systemic treatments were administered; however, the tumor was refractory, and the patient died 9 months after diagnosis. Conclusion: Prostate malignant lymphomas are rare but should be considered in patients with enlarged prostates and normal PSA levels. It should be noted that HIV patients have a high incidence of malignant lymphomas.

Type 1 diabetes mellitus after cord blood transplantation from an unrelated donor with a disease-sensitive haplotype.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT), a curative treatment for hematopoietic neoplasms, often causes various autoimmune disease-like conditions. In contrast, allo-HSCT-related type 1 diabetes mellitus is extremely rare. Herein, we report a case of allo-HSCT-related type 1 diabetes mellitus in a patient who had undergone cord blood transplantation (CBT) as a treatment for acute myeloid leukemia. The patient's human leukocyte antigen was replaced with the donor type after transplantation. The donor had a disease-sensitive haplotype. To the best of our knowledge, this is the first reported case of type 1 diabetes mellitus following CBT.