Cumulative autophagy insufficiency in mice leads to progression of ß-cell failure.

Autophagy is known to play a pivotal role in ß-cell function. While the lifelong inhibition of autophagy through Atg7 deletion in ß cells has been demonstrated to lead to impaired glucose tolerance together with ß-cell dysfunction, the temporal association between autophagy inhibition and ß-cell dysfunction remains unclear. To address such questions, inducible ß-cell-specific Atg7-knockout (ißAtg7KO) mice were generated, and autophagy inhibition was induced for two different time durations. Whereas 2 weeks of Atg7 ablation was sufficient to induce autophagy deficiency, confirmed by the accumulation of p62, ißAtg7KO mice exhibited normal glucose tolerance. In contrast, prolonged autophagy deficiency for 6 weeks resulted in glucose intolerance together with impaired insulin secretion. Direct mRNA sequencing and pathway analysis revealed that the gene set associated with insulin secretion was downregulated only after the 6-week prolonged autophagy inhibition. Furthermore, we identified a novel gene, Sprr1a, which was expressed at more than 50-fold higher levels during both the 2-week and 6-week autophagy inhibition. These findings suggest that autophagy insufficiency cumulatively leads to ß-cell failure after a certain interval, accompanied by stepwise alterations of gene expression patterns.

Efficient detection of copy-number variations using exome data: Batch- and sex-based analyses.

Many algorithms to detect copy number variations (CNVs) using exome sequencing (ES) data have been reported and evaluated on their sensitivity and specificity, reproducibility, and precision. However, operational optimization of such algorithms for a better performance has not been fully addressed. ES of 1199 samples including 763 patients with different disease profiles was performed. ES data were analyzed to detect CNVs by both the eXome Hidden Markov Model (XHMM) and modified Nord's method. To efficiently detect rare CNVs, we aimed to decrease sequencing biases by analyzing, at the same time, the data of all unrelated samples sequenced in the same flow cell as a batch, and to eliminate sex effects of X-linked CNVs by analyzing female and male sequences separately. We also applied several filtering steps for more efficient CNV selection. The average number of CNVs detected in one sample was <5. This optimization together with targeted CNV analysis by Nord's method identified pathogenic/likely pathogenic CNVs in 34 patients (4.5%, 34/763). In particular, among 142 patients with epilepsy, the current protocol detected clinically relevant CNVs in 19 (13.4%) patients, whereas the previous protocol identified them in only 14 (9.9%) patients. Thus, this batch-based XHMM analysis efficiently selected rare pathogenic CNVs in genetic diseases.

Conversion of pancreatic α cells into insulin-producing cells modulated by ß-cell insufficiency and supplemental insulin administration.

The emergence of bihormonal (BH) cells expressing insulin and glucagon has been reported under diabetic conditions in humans and mice. Whereas lineage tracing studies demonstrated that glucagon-producing α cells can be reprogrammed into BH cells, the underlying dynamics of the conversion process remain poorly understood. In the present study, we investigated the identities of pancreatic endocrine cells by genetic lineage tracing under diabetic conditions. When ß-cell ablation was induced by alloxan (ALX), a time-dependent increase in BH cells was subsequently observed. Lineage tracing experiments demonstrated that BH cells originate from α cells, but not from ß cells, in ALX-induced diabetic mice. Notably, supplemental insulin administration into diabetic mice resulted in a significant increase in α-cell-derived insulin-producing cells that did not express glucagon. Furthermore, lineage tracing in Ins2Akita diabetic mice demonstrated a significant induction of α-to-ß conversion. Thus, adult α cells have plasticity, which enables them to be reprogrammed into insulin-producing cells under diabetic conditions, and this can be modulated by supplemental insulin administration.

Heterogeneity of autophagic status in pancreatic ß cells under metabolic stress.

Autophagy in ß cells has been demonstrated to play a pivotal role in cellular homeostasis and the progression of glucose intolerance. Although autophagic activity is affected by metabolic stress both in vivo and in vitro, it remains unclear as to what extent the autophagic status in each ß cell is different from its neighboring cells. To address this question, GFP-LC3 reporter mice, which can visualize the autophagic status of each ß cell as green-fluorescent puncta, were crossed with obese diabetic db/db mice. Imaging of green-fluorescent puncta in the islets of GFP-LC3 mice revealed that ß cells are a heterogeneous population, as the density of GFP-LC3 puncta in each cell was variable. Furthermore, the variability was greater in GFP-LC3; db/db mice than in non-diabetic GFP-LC3; db/+ mice. Furthermore, when GFP-LC3 mice were treated with a low dose of S961, which antagonizes insulin signaling without inducing overt hyperglycemia, the number of ß cells with a high density of GFP puncta was increased, suggesting that insulin resistance affects autophagic status independently of glucose profiles. These results suggest that pancreatic ß cells under metabolic stress are heterogeneous regarding their autophagic status, which provides insights into the cellular dynamics of each ß cell rather than the whole ß-cell population.

A Case of Septic Pulmonary Embolism Caused by Pyelonephritis With Klebsiella pneumoniae in a Patient With Poorly Controlled Type 2 Diabetes Mellitus.

Septic pulmonary embolism (SPE) is caused by the microbe that is responsible for any clinical condition that may include urinary tract infections as in this case. We report a case of pyelonephritis with Klebsiella pneumoniae that led to SPE in an 80-year-old woman with poorly controlled diabetes mellitus (DM). Computed tomography (CT) revealed multiple nodules in the peripheral area of the bilateral lung and a contrast defect in the right renal vein, which was suspected to be an embolism. Blood and urine cultures revealed Klebsiella pneumoniae infection. These results confirmed the diagnosis of pyelonephritis and SPE. Treatment with ceftriaxone, cefazolin, and ciprofloxacin improved the patient's condition.

Two-year efficacy of lacosamide as adjunctive therapy for generalized tonic-clonic seizures in patients with juvenile myoclonic epilepsy.

OBJECTIVE: To report the long-term efficacy of adjunctive lacosamide therapy in patients with juvenile myoclonic epilepsy whose generalized tonic-clonic seizures were significantly reduced by treatment. METHODS: A retrospective study was conducted in patients who visited the Department of Child Neurology, National Hospital Organization Nishiniigata Chuo Hospital and the Department of Pediatrics, National Hospital Organization Nagasaki Medical Center. Among patients who had been diagnosed with juvenile myoclonic epilepsy, those who received lacosamide as adjunctive therapy for refractory generalized tonic-clonic seizures for at least 2 years from January 2017 to December 2022, and who achieved seizure freedom or >50% seizure reduction in tonic-clonic seizures were included. The medical records and neurophysiological data of the patients were reviewed retrospectively. RESULTS: Four patients met the inclusion criteria. The mean age at the onset of epilepsy was 11.3 years (range 10-12), and the mean age of starting lacosamide was 17.5 years (range 16-21). All patients received two or more antiseizure medications prior to lacosamide. Three of four patients had seizure freedom for more than 2 years, and the one remaining patient had >50% seizure reduction for more than one year. Only one patient had recurrent myoclonic seizures after starting lacosamide. The mean lacosamide dose at the last visit was 425 mg/day (range 300-600). CONCLUSION: Adjunctive lacosamide therapy might be a treatment option for juvenile myoclonic epilepsy with generalized tonic-clonic seizures, which are not responsive to standard antiseizure medications.

The HCN1 p.Ser399Pro variant causes epileptic encephalopathy with super-refractory status epilepticus.

HCN1 is one of four genes encoding hyperpolarization-activated cyclic nucleotide-gated channels. The phenotypic spectrum associated with HCN1 variants ranges from neonatal developmental and epileptic encephalopathy to idiopathic generalized epilepsy. We report a Japanese patient with repetitive focal seizures and super-refractory status epilepticus since early infancy caused by a de novo HCN1 variant, NM_021072.4, c.1195T>C, p.(Ser399Pro). This variant might have a dominant-negative effect on channel function, leading to severe epileptic encephalopathy.

STAT3 suppression and ß-cell ablation enhance α-to-ß reprogramming mediated by Pdx1.

As diabetes results from the absolute or relative deficiency of insulin secretion from pancreatic ß cells, possible methods to efficiently generate surrogate ß cells have attracted a lot of efforts. To date, insulin-producing cells have been generated from various differentiated cell types in the pancreas, such as acinar cells and α cells, by inducing defined transcription factors, such as PDX1 and MAFA, yet it is still challenging as to how surrogate ß cells can be efficiently generated for establishing future regenerative therapies for diabetes. In this study, we demonstrated that the exogenous expression of PDX1 activated STAT3 in α cells in vitro, and STAT3-null PDX1-expressing α cells in vivo resulted in efficient induction of α-to-ß reprogramming, accompanied by the emergence of α-cell-derived insulin-producing cells with silenced glucagon expression. Whereas ß-cell ablation by alloxan administration significantly increased the number of α-cell-derived insulin-producing cells by PDX1, STAT3 suppression resulted in no further increase in ß-cell neogenesis after ß-cell ablation. Thus, STAT3 modulation and ß-cell ablation nonadditively enhance α-to-ß reprogramming induced by PDX1, which may lead to the establishment of cell therapies for curing diabetes.

De novo ATP1A3 variants cause polymicrogyria.

Polymicrogyria is a common malformation of cortical development whose etiology remains elusive. We conducted whole-exome sequencing for 124 patients with polymicrogyria and identified de novo ATP1A3 variants in eight patients. Mutated ATP1A3 causes functional brain diseases, including alternating hemiplegia of childhood (AHC), rapid-onset dystonia parkinsonism (RDP), and cerebellar ataxia, areflexia, pes cavus, optic nerve atrophy, and sensorineural deafness (CAPOS). However, our patients showed no clinical features of AHC, RDP, or CAPOS and had a completely different phenotype: a severe form of polymicrogyria with epilepsy and developmental delay. Detected variants had different locations in ATP1A3 and different functional properties compared with AHC-, RDP-, or CAPOS-associated variants. In the developing cerebral cortex of mice, radial neuronal migration was impaired in neurons overexpressing the ATP1A3 variant of the most severe patients, suggesting that this variant is involved in cortical malformation pathogenesis. We propose a previously unidentified category of polymicrogyria associated with ATP1A3 abnormalities.

13q13.3 microdeletion associated with apparently balanced translocation of 46,XX,t(7;13) suggests NBEA involvement.

BACKGROUND: Deletion of 13q13.3 is an extremely rare event. CASE: We report on a 25-month-old girl with neurodevelopmental disorder and intellectual disability. She had dysmorphic facies characterized by synophrys, long and narrow palpebral fissures; and a large, round face with small organs such as the eyes and mouth positioned near the center. She was hypotonic and had autism-like behaviors. Blood tests and brain MRI revealed no specific findings. However, G-banding chromosome analysis showed an apparently balanced translocation: 46,XX,t(7,13)(q11.23;q12.3). Both parents had normal karyotypes. Furthermore, her abnormal phenotype and chromosomal breakpoint lesion were suspected to be associated. Hence, we conducted array comparative genomic hybridization, which revealed a 3.2 Mb novel pathological microdeletion at 13q13.3 involving 17 genes including neurobeachin (NBEA), a neurodevelopment disorder gene. Furthermore, fluorescence in situ hybridization using probes adjacent to the microdeletion suggested a concomitant occurrence of the deletion and translocation as the structural basis of this rare genomic variant. CONCLUSION: NBEA may have roles in her neurodevelopmental phenotypes, whereas other genes within the 13q13.3 microdeletion may contribute to her dysmorphic features.

Biphasic changes in ß-cell mass around parturition are accompanied by increased serotonin production.

Pancreatic ß-cell mass is known to be considerably altered during pregnancy and after parturition in rodents and humans. While ß-cell mass increases during pregnancy and starts to return toward its original level after parturition, the cellular mechanisms by which ß-cell mass during this period is regulated remains unclear. To address this issue in mice, we quantified ß-cell mass and investigated the mechanisms underlying its regulation throughout the perinatal and postpartum period. The increased ß-cell size and proliferation during pregnancy were significantly reduced shortly after parturition, whereas there was no evidence of ß-cell reprogramming or increased apoptosis. Direct RNA sequencing of islets from pregnant and postpartum mice demonstrated dynamic changes in gene expression patterns, showing robust downregulation of cell cycle-related genes 1 day after parturition, and the reupregulation of serotonin metabolism-related genes at postpartum day 7. Serotonin synthesis was activated only in lactating females, accompanied by increased ß-cell mass. Taken together, these findings demonstrate that ß-cell mass is decreased shortly after parturition owing to reduced ß-cell size and proliferation, and is subsequently increased, in association with lactation and serotonin biosynthesis.

[A case of advanced gastric cancer with disseminated carcinomatosis of bone marrow treated by S-1 and CDDP].

A 33-year-old man was admitted to our hospital due to DIC and multiple bone metastasis after distal gastrectomy for gastric cancer (Stage IIIB). We diagnosed disseminated carcinomatosis of bone marrow by gastric cancer. The patient was treated with combination chemotherapy of S-1 and CDDP (S-1 80 mg/m (2), po, day 1-21 and CDDP 60 mg/m(2), iv, day 8). After one course of the treatment, DIC was resolved and severe pain in his back and legs which had been poorly controlled was dramatically improved. He could thus be discharged from our hospital and survived for about six months. S-1 and CDDP therapy are considered to be effective for disseminated carcinomatosis of bone marrow due to gastric cancer, even if complicated by DIC.

Cellular Autophagy in α Cells Plays a Role in the Maintenance of Islet Architecture.

Autophagy is known to play a pivotal role in intracellular quality control through the degradation of subcellular damaged organelles and components. Whereas autophagy is essential for maintaining ß-cell function in pancreatic islets, it remains unclear as to how the cellular autophagy affects the homeostasis and function of glucagon-secreting α cells. To investigate the role of autophagy in α cells, we generated a mutant mouse model lacking Atg7, a key molecule for autophagosome formation, specifically in α cells. Histological analysis demonstrated more glucagon-positive cells, with a multilayered structure, in the islets under Atg7 deficiency, although metabolic profiles, such as body weight, blood glucose, and plasma glucagon levels were comparable between Atg7-deficient mice and control littermates. Consistent with our previous findings that Atg7 deficiency suppressed ß-cell proliferation, cellular proliferation was suppressed in Atg7-deficient α cells. These findings suggest that α-cell autophagy plays a role in maintaining α-cell area and normal islet architecture but appears to be dispensable for metabolic homeostasis.

Suppression of STAT3 signaling promotes cellular reprogramming into insulin-producing cells induced by defined transcription factors.

BACKGROUND: STAT3 has been demonstrated to play a role in maintaining cellular identities in the pancreas, whereas an activating STAT3 mutation has been linked to impaired ß-cell function. METHODS: The role of STAT3 in ß-cell neogenesis, induced by the exogenous expression of Pdx1, Neurog3, and Mafa, was analyzed in vitro and in vivo. FINDINGS: The expression of phosphorylated STAT3 (pSTAT3) was induced in both Pdx1-expressing and Mafa-expressing cells, but most of the induced ß cells were negative for pSTAT3. The suppression of STAT3 signaling, together with exogenously expressed Pdx1, Neurog3, and Mafa, significantly increased the number of reprogrammed ß cells in vitro and in vivo, enhanced the formation of islet-like clusters in mice, and ameliorated hyperglycemia in diabetic mice. INTERPRETATION: These findings suggest that STAT3 inhibition promotes cellular reprogramming into ß-like cells, orchestrated by defined transcription factors, which may lead to the establishment of cell therapies for curing diabetes. FUND: JSPS, MEXT, Takeda Science Foundation, Suzuken Memorial Foundation, Astellas Foundation for Research on Metabolic Disorders, Novo Nordisk, Eli Lilly, MSD, Life Scan, Novartis, and Takeda.

Current Therapeutic Drugs Against Cerebral Vasospasm after Subarachnoid Hemorrhage: A Comprehensive Review of Basic and Clinical Studies.

BACKGROUND: Cerebral vasospasm (CVS) is well known as a major complication in subarachnoid hemorrhage (SAH) patients, and research has long been focused on improving morbidity and mortality. As CVS commonly develops from day 4 to day 14 after SAH onset, SAH patients require therapies with drugs for preventing CVS after surgical treatment for the source of hemorrhage, mostly ruptured intracranial aneurysms. It is thought that the pathogenesis of CVS is initiated by prolonged smooth muscle contraction, and the subsequent hypoperfusion and cytotoxic responses induce cerebral ischemia. Although therapeutic investigations have historically focused on morphological improvement, the improvement of outcome is limited by the reversal of arterial narrowing. Therefore, it might be important to look back at evidence from long-lasting studies of CVS and to determine a highroad to effective drugs, including combination therapy. OBJECTIVE: In this review, we introduce current candidate beneficial drugs against CVS in clinical SAH, including nimodipine and other Ca2+ channel antagonists, magnesium sulfate, clazosentan, statins, cilostazol, eicosapentaenoic acid, fasudil hydrochloride, milrinone, and edaravone, all of which have been frequently studied in recent years.

Sequential changes in Rotterdam CT scores related to outcomes for patients with traumatic brain injury who undergo decompressive craniectomy.

OBJECT Rotterdam CT scoring is a CT classification system for grouping patients with traumatic brain injury (TBI) based on multiple CT characteristics. This retrospective study aimed to determine the relationship between initial or preoperative Rotterdam CT scores and TBI prognosis after decompressive craniectomy (DC). METHODS The authors retrospectively reviewed the medical records of all consecutive patients who underwent DC for nonpenetrating TBI in 2 hospitals from January 2006 through December 2013. Univariate and multivariate logistic regression and receiver operating characteristic (ROC) curve analyses were used to determine the relationship between initial or preoperative Rotterdam CT scores and mortality at 30 days or Glasgow Outcome Scale (GOS) scores at least 3 months after the time of injury. Unfavorable outcomes were GOS Scores 1-3 and favorable outcomes were GOS Scores 4 and 5. RESULTS A total of 48 cases involving patients who underwent DC for TBI were included in this study. Univariate analyses showed that initial Rotterdam CT scores were significantly associated with mortality and both initial and preoperative Rotterdam CT scores were significantly associated with unfavorable outcomes. Multivariable logistic regression analysis adjusted for established predictors of TBI outcomes showed that initial Rotterdam CT scores were significantly associated with mortality (OR 4.98, 95% CI 1.40-17.78, p = 0.01) and unfavorable outcomes (OR 3.66, 95% CI 1.29-10.39, p = 0.02) and preoperative Rotterdam CT scores were significantly associated with unfavorable outcomes (OR 15.29, 95% CI 2.50-93.53, p = 0.003). ROC curve analyses showed cutoff values for the initial Rotterdam CT score of 5.5 (area under the curve [AUC] 0.74, 95% CI 0.59-0.90, p = 0.009, sensitivity 50.0%, and specificity 88.2%) for mortality and 4.5 (AUC 0.71, 95% CI 0.56-0.86, p = 0.02, sensitivity 62.5%, and specificity 75.0%) for an unfavorable outcome and a cutoff value for the preoperative Rotterdam CT score of 4.5 (AUC 0.81, 95% CI 0.69-0.94, p < 0.001, sensitivity 90.6%, and specificity 56.2%) for an unfavorable outcome. CONCLUSIONS Assessment of changes in Rotterdam CT scores over time may serve as a prognostic indicator in TBI and can help determine which patients require DC.

Bilateral posterior fossa chronic subdural hematoma treated with craniectomy: Case report and review of the literature.

BACKGROUND: Posterior chronic subdural hematomas (pCSHs) are rare. Their diagnosis and treatment are difficult. DESCRIPTION: A 69-year-old woman was admitted to our hospital with nausea, headache, and mild consciousness disturbance. Computed tomography and magnetic resonance imaging showed bilateral pCSH. To prevent further neurological deterioration, we performed surgery under general anesthesia by midline suboccipital craniectomy. Unexpected bleeding from a developed circuitous occipital sinus was stopped with hemoclips. After hematoma removal, she recovered and was transferred to a rehabilitation hospital. By the 19(th) postoperative day, she had developed no neurologic deficits. CONCLUSION: This experience demonstrates the risk of blind surgical therapy in patients with pCSH. In such patients, posterior fossa craniectomy may be preferable in terms of diagnosis and safe treatment.

A spontaneous superficial temporal artery pseudoaneurysm possibly related to atherosclerosis: Case report and review of literature.

BACKGROUND: Spontaneous superficial temporal artery (STA) pseudoaneurysms are very rare; only four cases, including ours, have been reported to date. Therefore, the cause of them has not been studied. CASE DESCRIPTION: A 57-year-old woman was admitted to our hospital with a pulsatile mass in the left preauricular region. Her medical history included hypertension, dyslipidemia, and angina pectoris. She denied a history of head injury or minor head trauma. Three-dimensional computed tomography angiography showed a well-enhanced saccular aneurysm on the main trunk of the STA. To prevent rupture it was removed surgically. The histological diagnosis was pseudoaneurysm with atherosclerosis. By the 2(nd) postoperative day, she had completely recovered and was discharged home. There has been no relapse. CONCLUSIONS: As all four documented patients were at high risk for atherosclerosis, we posit that a causal factor was weakening of the arterial wall due to atherosclerosis and chronic pressure on the STA from anatomical structures. Here, we present histological evidence to support this hypothesis.

Brain metastases from asymptomatic adenocarcinoma of the pancreas: an autopsy case report.

BACKGROUND: Brain metastasis from pancreatic cancer is extremely rare. Because pancreatic cancer usually has a rapidly progressive nature, the majority of affected patients die from primary lesions before exhibiting clinical signs suggestive of brain metastases. CASE DESCRIPTION: The patient was a 62-year-old man who developed generalized convulsion followed by right hemiparesis accompanied by decreased consciousness level. Computed tomography (CT) scan revealed multiple brain tumors with ring-like contrast enhancement. Stereotactic biopsy disclosed mucinous adenocarcinoma. A marked increase in the serum CA19-9 level was noted, but the patient did not exhibit any other signs of pancreatic disease. Repeated whole body CT scan and ultrasonography demonstrated no primary lesions. The patient died of multi-organ failure during chemotherapy combined with radiation for metastatic brain tumors. Autopsy revealed well-differentiated papillary adenocarcinoma in the pancreatic head and systemic metastases associated with tumor emboli were widely distributed in various organs. CONCLUSION: This patient initially presented only with symptoms of neurologic disorder and no pancreatic symptoms. Moreover, repeated radiologic examinations did not reveal the primary lesion. We considered that the unusual clinical course in our patient may be partly explained by the autopsy findings: diffuse sclerotic changes of the pancreas without swelling. The present report suggests that undetected pancreatic cancer may have been the primary lesion classified as "unknown origin" in some cases of metastatic brain tumors.