Use of bispectral index for detection of partial cerebral hypoperfusion during cervical spine surgery: A case report.

The BIS value may decrease by cerebral hypoperfusion. We report a case in which the BIS value suddenly decreased during cervical spine surgery, which led us to find cervical screws compressing the vertebral arteries. In a 79-year-old man undergoing cervical spine surgery, the BIS suddenly decreased from about 40 to 10-20, about 4 h after the start of surgery. Intraoperative 3-dementional computed tomography indicated that both the two tips of cervical screws inserted in the 6th cervical vertebra were within bilateral transverse foramens. These cervical screws were removed, and the BIS increased immediately. The cervical screws were re-inserted again thorough the same vertebra into the bilateral transverse foramens, and the BIS decreased immediately. Postoperatively, cerebral hypoperfusion due to compression of bilateral vertebral arteries by two cervical screws was identified. The BIS may be a useful to detect cerebral hypoperfusion due to compression of the vertebral artery by a cervical screw.

Effect of Recent Antirheumatic Drug on Features of Rheumatoid Arthritis-Associated Lymphoproliferative Disorders.

OBJECTIVE: In this study, we examine how advancements in novel antirheumatic drugs affect the clinicopathologic features of lymphoproliferative disorder (LPD) in patients with rheumatoid arthritis (RA). METHODS: In this multicenter study across 53 hospitals in Japan, we characterized patients with RA who developed LPDs and visited the hospitals between January 1999 and March 2021. The statistical tools used included Fisher's exact test, the Mann-Whitney U-test, the log-rank test, logistic regression analysis, and Cox proportional hazards models. RESULTS: Overall, 752 patients with RA-associated LPD (RA-LPD) and 770 with sporadic LPD were included in the study. We observed significant differences in the clinicopathologic features between patients with RA-LPD and those with sporadic LPD. Histopathological analysis revealed a high frequency of LPD-associated immunosuppressive conditions. Furthermore, patients with RA-LPD were evaluated based on the antirheumatic drugs administered. The methotrexate (MTX) plus tacrolimus and MTX plus tumor necrosis factor inhibitor (TNFi) groups had different affected site frequencies and histologic subtypes than the MTX-only group. Moreover, MTX and TNFi may synergistically affect susceptibility to Epstein-Barr virus infection. In case of antirheumatic drugs administered after LPD onset, tocilizumab (TCZ)-only therapy was associated with lower frequency of regrowth after spontaneous regression than other regimens. CONCLUSION: Antirheumatic drugs administered before LPD onset may influence the clinicopathologic features of RA-LPD, with patterns changing over time. Furthermore, TCZ-only regimens are recommended after LPD onset.

Horizontal and vertical microchamber platforms for evaluation of the paracellular permeability of an epithelial cell monolayer.

Epithelial cells serve as a barrier by tightly adhering to each other and contribute to the homeostasis of living organisms by controlling substance permeation. Therefore, evaluation of the barrier function is important in pharmaceutical development processes. However, the widely used Transwell-based assays require the development of the defect-free epithelial cell monolayer above several tens of mm2, often resulting in low reproducibility and requiring a long incubation time. In addition, the culture surface of cells is far from the bottom of the well plate, making it difficult to observe the cell morphology using an optical microscope. Herein, we propose simple polydimethylsiloxane microfluidic devices for evaluating the barrier function of an epithelial monolayer using a microchamber array. After the formation of the epithelial monolayer over microchambers, the permeation of the marker molecules introduced above resulted in increased fluorescence intensity in microchambers, which was monitored using confocal laser scanning microscopy. We show that using this technique, alteration of the paracellular permeability induced by sodium caprate (C10) and cytochalasin-D, permeation enhancing factors, can be elucidated. Furthermore, by tilting the microchamber device 90 degrees, the vertical cell section and microchambers were imaged in the same focal plane, allowing for live visualization of the passage of fluorescent substances across the cell monolayer. This technique is expected to be useful for investigating the relationship between paracellular permeability and cell morphology, which is unattainable through conventional methods.

Cooperative contributions of the klf1 and klf17 genes in zebrafish primitive erythropoiesis.

Krüppel-like transcription factors (Klfs), which are characterized by the three conserved C-terminal zinc fingers, are involved in various biological processes, such as haematopoiesis and angiogenesis. However, how the Klf family of transcription factors cooperate in organogenesis remains elusive. During zebrafish embryogenesis, both klf1 and klf17 are expressed in the intermediate cell mass (ICM), where primitive erythroid cells are produced. Using CRISPR-Cas9 genome editing technology, we established klf1-klf17 double mutant zebrafish to investigate the functionally interactive roles of the klf1 and klf17 genes. The klf1-klf17 mutant exhibited a diminished number of circulating primitive erythroid cells at 2 days postfertilization (dpf), while klf1 or klf17 single mutants and wild-type embryos produced comparable numbers of primitive erythroid cells. Circulating erythroid cells from the klf1-klf17 mutant possessed larger nuclei at 2 dpf than wild-type cells, suggesting the impairment of primitive erythroid cell maturation. The expression of the erythroid cell maturation markers band3 and mitoferrin, but not the haematopoietic progenitor markers c-myb and scl, was decreased in the klf1-klf17 mutant at 1 dpf. Thus, these results illustrate the cooperative function of klf1 and klf17 in the maturation processes of zebrafish primitive erythroid cells.

Identification of key microRNAs regulating ELOVL6 and glioblastoma tumorigenesis.

ELOVL fatty acid elongase 6 (ELOVL6) controls cellular fatty acid (FA) composition by catalyzing the elongation of palmitate (C16:0) to stearate (C18:0) and palmitoleate (C16:1n-7) to vaccinate (C18:1n-7). Although the transcriptional regulation of ELOVL6 has been well studied, the post-transcriptional regulation of ELOVL6 is not fully understood. Therefore, this study aims to evaluate the role of microRNAs (miRNAs) in regulating human ELOVL6. Bioinformatic analysis identified five putative miRNAs: miR-135b-5p, miR-135a-5p, miR-125a-5p, miR-125b-5p, and miR-22-3p, which potentially bind ELOVL6 3'-untranslated region (UTR). Results from dual-luciferase assays revealed that these miRNAs downregulate ELOVL6 by directly interacting with the 3'-UTR of ELOVL6 mRNA. Moreover, miR-135b-5p and miR-135a-5p suppress cell proliferation and migration in glioblastoma multiforme cells by inhibiting ELOVL6 at the mRNA and protein levels. Taken together, our results provide novel regulatory mechanisms for ELOVL6 at the post-transcriptional level and identify potential candidates for the treatment of patients with glioblastoma multiforme.

Takotsubo cardiomyopathy with left ventricle thrombus caused by subacute thyroiditis.

We report a rare case of takotsubo cardiomyopathy caused by subacute thyroiditis in a man in his 50s. He went to the doctor with complaints of loss of appetite, diarrhoea, chills and general malaise. He had consciousness disturbance, thyrotoxicosis and thyroid-stimulating hormone (TSH) suppression. Thyroglobulin and C reactive protein levels in the blood were elevated, but TSH receptor antibody, thyroid-stimulating antibody, antithyroglobulin antibody and antithyroid peroxidase antibody were not. We began treatment with prednisolone and propranolol after he was diagnosed with thyroid storm caused by subacute thyroiditis. The ECG revealed inverted T waves on the fifth day after admission. He was newly diagnosed with takotsubo cardiomyopathy on the day. A large thrombus was detected in the left ventricle, requiring anticoagulation therapy. Thus, even if there are no findings of takotsubo cardiomyopathy or thrombus at the onset of thyroid storm, appropriate monitoring is required because they can develop during the treatment course.

Programmed death ligand 1-positive immune cells in primary tumor or metastatic axillary lymph nodes can predict prognosis of triple-negative breast cancer even when present at < 1% in the tumor region.

BACKGROUND: The efficacy of pre-operative systemic treatment (PST) combined with immune checkpoint inhibition (ICI) for triple-negative breast cancer (TNBC) has been recognized recently as being independent of the degree of programmed death ligand-1 (PD-L1) positivity of infiltrating immune cells, especially for patients with axillary lymph node metastasis (ALNM). METHODS: TNBC patients with ALNM were treated surgically between 2002 and 2016 in our facility (n = 109), of whom 38 received PST before resection. The presence of tumor-infiltrating lymphocytes (TILs) expressing CD3, CD8, CD68, PD-L1 (detected by antibody SP142) and FOXP3 at primary and metastatic LN sites was quantified. RESULTS: The size of invasive tumor and the number of metastatic axillary LN were confirmed as prognostic markers. The numbers of both CD8+ and FOXP3+ TILs at primary sites were also recognized as prognostic markers, especially for overall survival (OS) (CD8, p = 0.026; FOXP3, p < 0.001). The presence of CD8+, FOXP3+ and PD-L1+ cells was better maintained in LN after PST and may contribute to improved antitumor immunity. Provided they were present as clusters of ≥ 70 positive cells, even < 1% of immune cells expressing PD-L1 at primary sites predicted a more favorable prognosis for both disease-free survival (DFS) (p = 0.004) and OS (p = 0.020). This was the case not only for 30 matched surgical patients, but also in all 71 surgical only patients (DFS: p < 0.001 and OS: p = 0.002). CONCLUSIONS: PD-L1+ , CD8+ or FOXP3+ immune cells in the tumor microenvironment (TME) at both primary and metastatic sites are significant on prognosis, which could be a clue to expect the potential for better responses to the combination of chemotherapy and ICI, especially for patients with ALNM.

L-Asparaginase-Induced Continuous Hyperglycemia With Type 1 Diabetes-Related Antibodies and HLA Genotypes: A Case Study.

A 19-year-old male presented with fatigue and dyspnea on exertion. He was diagnosed with acute T-cell lymphoblastic leukemia. After following the Group for Research on Adult Acute Lymphoblastic Leukemia (GRAALL) 2003 protocol that incorporates L-asparaginase (L-Asp) treatment, blood glucose levels became elevated for more than one year and insulin secretion was depleted. Anti-glutamic acid decarboxylase (GAD) and anti-islet antigen 2 (IA-2) antibody levels were both positive, which is rare. The patient's HLA genotype was sensitive for type 1 diabetes. L-Asp can cause transient hyperglycemia as a side effect. However, cases with the anti-GAD antibody have not been reported in L-Asp-induced diabetes. In summary, L-Asp-induced continuous hyperglycemia might be associated with a type 1 diabetes-related HLA genotype through elevations of anti-GAD and anti-IA-2 antibodies.

Controlling Nutritional Status (CONUT) Score and Sarcopenia as Mutually Independent Prognostic Biomarkers in Advanced Urothelial Carcinoma.

Background: While the controlling nutritional status (CONUT) score and sarcopenia are objective indices of different aspects of a patient's general condition, few studies have comprehensively examined their mutual relationship in patients with advanced cancer. Methods: This retrospective study included 200 Japanese patients with advanced urothelial carcinoma (aUC). Sarcopenia was diagnosed using Prado's definition. The CONUT score and sarcopenia were examined for their possible association, and their prognostic value was analyzed. Results: The CONUT score and sarcopenia were not significantly associated. While sarcopenia occurred in 168 patients (84%), more than half of them had normal or only slightly impaired nutritional status, as indicated by a CONUT score of 0−2. During follow-up (median: 13.3 months), 149 patients died. The CONUT score and sarcopenia were independent prognostic factors (hazard ratio 1.22 and 2.23, respectively; both p < 0.001), whereas performance status was not. Incorporating the CONUT score, sarcopenia, and both into Bajorin's and Apolo's prognostic models increased their concordance index as follows: 0.612 for Bajorin's original model to 0.653 (+the CONUT score), 0.631 (+sarcopenia), and 0.665 (+both), and 0.634 for Apolo's original model to 0.655 (+the CONUT score), 0.653 (+ sarcopenia), and 0.668 (+both). Conclusion: The CONUT score and sarcopenia were mutually independent in terms of their prognostic value in patients with aUC. These objective indices of a patient's general condition may help in decision-making when considering treatment for patients with aUC.

ILC3s select microbiota-specific regulatory T cells to establish tolerance in the gut.

Microbial colonization of the mammalian intestine elicits inflammatory or tolerogenic T cell responses, but the mechanisms controlling these distinct outcomes remain poorly understood, and accumulating evidence indicates that aberrant immunity to intestinal microbiota is causally associated with infectious, inflammatory and malignant diseases1-8. Here we define a critical pathway controlling the fate of inflammatory versus tolerogenic T cells that respond to the microbiota and express the transcription factor RORγt. We profiled all RORγt+ immune cells at single-cell resolution from the intestine-draining lymph nodes of mice and reveal a dominant presence of T regulatory (Treg) cells and lymphoid tissue inducer-like group 3 innate lymphoid cells (ILC3s), which co-localize at interfollicular regions. These ILC3s are distinct from extrathymic AIRE-expressing cells, abundantly express major histocompatibility complex class II, and are necessary and sufficient to promote microbiota-specific RORγt+ Treg cells and prevent their expansion as inflammatory T helper 17 cells. This occurs through ILC3-mediated antigen presentation, αV integrin and competition for interleukin-2. Finally, single-cell analyses suggest that interactions between ILC3s and RORγt+ Treg cells are impaired in inflammatory bowel disease. Our results define a paradigm whereby ILC3s select for antigen-specific RORγt+ Treg cells, and against T helper 17 cells, to establish immune tolerance to the microbiota and intestinal health.

Salivary gland polymorphous adenocarcinoma: Clinicopathological features and gene alterations in 36 Japanese patients.

BACKGROUND: Polymorphous adenocarcinoma is a common intraoral minor salivary gland carcinoma in Western countries but is extremely rare in Japan. The current study aimed to characterize the clinicopathological features and status of molecular alterations of polymorphous adenocarcinoma-associated genes, such as PRKD1/2/3, ARID1A, and DDX3X, in a large cohort of Japanese patients with polymorphous adenocarcinoma. METHODS: We examined the cases of 36 Japanese patients with salivary gland polymorphous adenocarcinoma and 26 cases involving histopathological mimics. To detect gene splits, fluorescence in situ hybridization was carried out for polymorphous adenocarcinoma-associated genes. Additionally, we applied a SNaPshot multiplex assay to identify PRKD1 hotspot mutations. RESULTS: This study revealed the indolent clinical course of polymorphous adenocarcinoma with a high 10-year overall survival rate (92.9%), accompanied by occasional local recurrences and cervical lymph node metastasis (23.3%). Twenty cases (55.6%) of polymorphous adenocarcinoma (but none of the mimics) exhibited alterations in at least one polymorphous adenocarcinoma-associated gene. Rearrangement of polymorphous adenocarcinoma-associated genes and PRKD1 E710D were identified in 17 (47.2%) and 4 (11.1%) cases, respectively; one case showed coexisting PRKD3 split and PRKD1 E710D. In the multivariate analysis, high clinical stage (p = 0.0005), the presence of prominent nucleoli (p = 0.0003), and ARID1A split positivity (p = 0.004) were independent risk factors for disease-free survival. CONCLUSION: Japanese patients with polymorphous adenocarcinoma showed clinicopathological features similar to those reported in Western countries. This study disclosed that polymorphous adenocarcinoma-associated genetic alterations were common and specific findings in polymorphous adenocarcinomas. The diagnostic role and possible prognostic significance of polymorphous adenocarcinoma-associated genetic alterations in polymorphous adenocarcinomas were suggested.

Initial screening by immunohistochemistry is effective in universal screening for Lynch syndrome in endometrial cancer patients: a prospective observational study.

BACKGROUND: Few prospective reports of universal screening for Lynch syndrome exist for patients with endometrial cancer. In this study, we performed immunohistochemical staining for DNA mismatch repair-related genes (MLH1, MSH2, MSH6 and PMS2), to determine the extent to which Lynch syndrome can be diagnosed in endometrial cancer patients through universal screening. METHODS: We recruited 116 consecutive patients assumed to have uterine corpus malignancy from October 2019 to February 2021 in a prospective observational study. We performed immunohistochemical for mismatch repair-related proteins on samples from 100 patients who had surgicopathologically confirmed diagnoses of endometrial cancer. Samples with missing immunohistochemical results for any of the proteins had subsequent universal screening tests for microsatellite instability, DNA methylation of the MLH1 promoter region and mismatch repair genetics. RESULTS: We identified 19 (19.0%) patients with lost results for any of the proteins. All 19 patient samples had subsequent screening tests. We identified the microsatellite instability-high phenotype in 84.2% (16/19) of these patients and MLH1 methylation in 57.9% (11/19). Mismatch repair genetic testing detected two pathological variants, in MSH2 and MSH6, which indicated that the prevalence of Lynch syndrome was 2.0% in our cohort. Two cases of unclassified variant (MSH6) and one case of benign variant (PMS2) were also detected. CONCLUSIONS: Initial screening by immunohistochemical is an effective method in universal screening for Lynch syndrome in endometrial cancer patients.

Preoperative models incorporating the systemic immune-inflammation index for predicting prognosis and muscle invasion in patients with non-metastatic upper tract urothelial carcinoma.

OBJECTIVE: To develop preoperative models as a guide to indications for neoadjuvant chemotherapy (NAC) and regional lymph node dissection (LND) before and at radical nephroureterectomy (RNU), respectively, in patients with non-metastatic upper tract urothelial carcinoma (UTUC) by incorporating the systemic immune-inflammation index (SII). METHODS: This retrospective study enrolled 103 consecutive patients with UTUC undergoing RNU. The SII was calculated as neutrophils × platelets / lymphocytes. Multivariable Cox proportional hazard model was used to develop preoperative models for cancer-specific survival (CSS) and overall survival (OS). A model for predicting muscle invasion was developed using logistic regression analysis. Harrell's concordance-index (c-index) or the area under the receiver operating characteristic curve (AUC) was used to evaluate the accuracy of the models. RESULTS: During follow-up (median: 41 months), 26 and three patients died of UTUC and other causes, respectively. Performance status > 0, clinical tumor (cT) stage ≥ 3, and SII > 520 were independent adverse prognosticators for CSS, and one point was assigned to each prognosticator. Risk score models comprising the sum of the points stratified patients into three risk groups (0, 1, and 2-3; P < 0.001 for CSS and OS) with respective c-indices of 0.843 and 0.820. SII > 677 and ≥ cT3 were independently associated with muscle invasion. A model based on these variables predicted muscle invasion with AUC of 0.804. CONCLUSION: Preoperative SII is significantly associated with worse survival outcomes and muscle invasion in patients with non-metastatic UTUC. Our preoperative predictive models may serve as a guide to indications for NAC and LND.

Dysregulation of ILC3s unleashes progression and immunotherapy resistance in colon cancer.

Group 3 innate lymphoid cells (ILC3s) regulate immunity and inflammation, yet their role in cancer remains elusive. Here, we identify that colorectal cancer (CRC) manifests with altered ILC3s that are characterized by reduced frequencies, increased plasticity, and an imbalance with T cells. We evaluated the consequences of these changes in mice and determined that a dialog between ILC3s and T cells via major histocompatibility complex class II (MHCII) is necessary to support colonization with microbiota that subsequently induce type-1 immunity in the intestine and tumor microenvironment. As a result, mice lacking ILC3-specific MHCII develop invasive CRC and resistance to anti-PD-1 immunotherapy. Finally, humans with dysregulated intestinal ILC3s harbor microbiota that fail to induce type-1 immunity and immunotherapy responsiveness when transferred to mice. Collectively, these data define a protective role for ILC3s in cancer and indicate that their inherent disruption in CRC drives dysfunctional adaptive immunity, tumor progression, and immunotherapy resistance.

Simple stain-free screening method for pectinolytic microorganisms under alkalophilic conditions.

OBJECTIVES: To develop a simple pectin-degrading microorganism screening method. RESULTS: We developed a method utilizing the phenomenon whereby cooling an alkaline agar medium containing pectin causes the agar to become cloudy. This highly simplified method involves culturing the microorganisms on pectin-containing agar medium until colony formation is observed, and subsequent overnight cooling of the agar medium to 4 °C. Using this simple procedure, we successfully identified pectin-degrading microorganisms by observing colonies with halos on the clouded agar medium. We used alkaline pectinase and Bacillus halodurans, which is known to secrete alkaline pectinase, to establish the screening method. We demonstrated the screening of pectin-degrading microorganisms using the developed method and successfully isolated pectin-degrading microorganisms (Paenibacillus sp., Bacillus clausii, and Bacillus halodurans) from a soil sample. CONCLUSIONS: The developed method is useful for identifying pectin-degrading microorganisms.

Prognostic significance of the controlling nutritional status score in patients with metastatic renal cell carcinoma diagnosed before an era of first-line immune-oncology combination therapies.

OBJECTIVE: To explore the prognostic role of the controlling nutritional status score in patients with metastatic renal cell carcinoma. METHODS: We retrospectively analyzed 107 patients with metastatic renal cell carcinoma who received their diagnosis between 2007 and 2018 and were treated with or without a first-line interferon or tyrosine kinase inhibitor at a single cancer center. The controlling nutritional status score was based on values for albumin, lymphocyte count and total cholesterol at the metastatic renal cell carcinoma diagnosis. Association of the controlling nutritional status score and clinical variables, including the Memorial Sloan-Kettering Cancer Center and the International Metastatic Renal Cell Carcinoma Database Consortium risk classifications, with overall survival was examined using the Cox proportional hazard model. Predictive accuracy of the prognostic factors was assessed using Harrell's concordance index. RESULTS: First-line interferon and tyrosine kinase inhibitor were given to 48 (45%) and 41 (38%) patients, respectively, and 28 (26%) and 33 (31%) patients underwent cytoreductive nephrectomy and metastasectomy, respectively. During follow-up (median: 36.3 months), 64 patients died. The median controlling nutritional status score was 2 (range: 0-8). A controlling nutritional status score ≥ 2 was significantly associated with shorter overall survival (P < 0.01) independently of the Memorial Sloan-Kettering Cancer Center and International Metastatic Renal Cell Carcinoma Database Consortium risk classifications. Integration of the controlling nutritional status score into the Memorial Sloan-Kettering Cancer Center and International Metastatic Renal Cell Carcinoma Database Consortium risk classifications improved concordance index from 0.702 to 0.770 and from 0.698 to 0.749, respectively. CONCLUSION: The controlling nutritional status score may serve as a prognostic biomarker objectively reflecting the general physical condition of patients with metastatic renal cell carcinoma treated with or without first-line interferon or tyrosine kinase inhibitor in terms of nutritional and immuno-inflammatory status.

Proline-arginine poly-dipeptide encoded by the C9orf72 repeat expansion inhibits adenosine deaminase acting on RNA.

A GGGGCC hexanucleotide repeat expansion in the C9orf72 gene is linked to the pathogenesis of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) (C9-ALS/FTD). Unconventional translation of the hexanucleotide repeat expansion generates five dipeptide repeat proteins (DPRs). The molecular mechanism underlying the DPR-linked neurotoxicity is under investigation. In this study, using cell-based models, we show that poly-proline-arginine DPR (poly-PR), the most neurotoxic DPR in vitro, binds to adenosine deaminase acting on RNA (ADAR)1p110 and ADAR2 and inhibits their RNA editing activity. We further show that poly-PR impairs cellular stress response that is mediated by ADAR1p110. These results together suggest that the poly-PR-mediated inhibition of the ADAR activity contributes to C9-ALS/FTD-linked neurotoxicity.

Optimal timing of tracheostomy in patients on veno-venous extracorporeal membrane oxygenation for coronavirus 2019: a case series.

AIM: An early tracheostomy is often considered for patients with veno-venous extracorporeal membrane oxygenation (VV-ECMO). However, there is no consensus on the timing of a tracheostomy in patients on VV-ECMO for coronavirus disease 2019 (COVID-19). The present report described the optimal timing of tracheostomy for these patients. METHOD: The present study was a single-center case series. We retrospectively reviewed the medical records of nine consecutive patients who underwent tracheostomy either during or after VV-ECMO treatment in our center between January 1, 2020 and December 31, 2020. RESULTS: All the patients received a percutaneous dilatational tracheostomy, which was performed during VV-ECMO in four patients. Three of these patients experienced hemorrhagic complications, and the remaining patient required a circuit change on the day after the operation. Heparin was discontinued 8 h preoperatively and resumed 1-14 h later. The platelet count was below normal in two patients, but no transfusion was performed. APTT was almost normal, and D-dimer was elevated postoperatively. The remaining five patients received a tracheostomy after weaning off VV-ECMO, and no complication was observed. Eight patients were deeply sedated during VV-ECMO to prioritize lung rest and prevent infecting the healthcare workers. CONCLUSION: In the present study, patients who underwent a tracheostomy during VV-ECMO tended to have more hemorrhagic complications. Because an early tracheostomy during ECMO has little benefit for patients with COVID-19, it should be performed after weaning off VV-ECMO to protect the safety of the healthcare workers concerned.