Structural basis for ion selectivity in potassium-selective channelrhodopsins.

KCR channelrhodopsins (K+-selective light-gated ion channels) have received attention as potential inhibitory optogenetic tools but more broadly pose a fundamental mystery regarding how their K+ selectivity is achieved. Here, we present 2.5-2.7 Å cryo-electron microscopy structures of HcKCR1 and HcKCR2 and of a structure-guided mutant with enhanced K+ selectivity. Structural, electrophysiological, computational, spectroscopic, and biochemical analyses reveal a distinctive mechanism for K+ selectivity; rather than forming the symmetrical filter of canonical K+ channels achieving both selectivity and dehydration, instead, three extracellular-vestibule residues within each monomer form a flexible asymmetric selectivity gate, while a distinct dehydration pathway extends intracellularly. Structural comparisons reveal a retinal-binding pocket that induces retinal rotation (accounting for HcKCR1/HcKCR2 spectral differences), and design of corresponding KCR variants with increased K+ selectivity (KALI-1/KALI-2) provides key advantages for optogenetic inhibition in vitro and in vivo. Thus, discovery of a mechanism for ion-channel K+ selectivity also provides a framework for next-generation optogenetics.

Structural basis for channel conduction in the pump-like channelrhodopsin ChRmine.

ChRmine, a recently discovered pump-like cation-conducting channelrhodopsin, exhibits puzzling properties (large photocurrents, red-shifted spectrum, and extreme light sensitivity) that have created new opportunities in optogenetics. ChRmine and its homologs function as ion channels but, by primary sequence, more closely resemble ion pump rhodopsins; mechanisms for passive channel conduction in this family have remained mysterious. Here, we present the 2.0 Å resolution cryo-EM structure of ChRmine, revealing architectural features atypical for channelrhodopsins: trimeric assembly, a short transmembrane-helix 3, a twisting extracellular-loop 1, large vestibules within the monomer, and an opening at the trimer interface. We applied this structure to design three proteins (rsChRmine and hsChRmine, conferring further red-shifted and high-speed properties, respectively, and frChRmine, combining faster and more red-shifted performance) suitable for fundamental neuroscience opportunities. These results illuminate the conduction and gating of pump-like channelrhodopsins and point the way toward further structure-guided creation of channelrhodopsins for applications across biology.

Needle tract seeding following endoscopic ultrasound-guided fine-needle aspiration for pancreatic cancer: a report of two cases.

BACKGROUND: Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) is a useful tool in pancreatic cancer diagnosis. However, the procedure itself may cause peritoneal dissemination and needle tract seeding at the puncture site. We herein report two cases of gastric wall metastasis due to needle tract seeding after EUS-FNA. CASE PRESENTATION: Case 1: A 68-year-old woman was admitted to our hospital for persistent cough. Computed tomography (CT) scan revealed inflammatory changes in the left lung field, and incidentally, a 15-mm hypovascular mass was detected in the pancreatic body. She underwent EUS-FNA and was diagnosed as pancreatic adenocarcinoma. She underwent distal pancreatectomy with splenectomy; however, a small hard mass was observed in the posterior gastric wall during surgery. We performed partial gastrectomy, and the resected specimen was diagnosed as a needle tract seeding following EUS-FNA. She then underwent adjuvant chemotherapy with TS-1, but the pancreatic cancer showed recurrence 6 months after surgery. She died due to peritoneal dissemination 18 months after surgery. Case 2: A 70-year-old man was incidentally detected with a pancreatic body mass on a CT scan as part of his follow-up for recurrence of basal cell carcinoma. He underwent EUS-FNA and was diagnosed as pancreatic adenocarcinoma. He had nodules in both lungs, and it was difficult to differentiate them from lung metastasis of pancreatic cancer. Therefore, he underwent neoadjuvant chemoradiotherapy, and thereafter, the lung nodules showed no changes; hence, he underwent distal pancreatectomy with splenectomy. During surgery, we observed a hard mass in the posterior gastric wall. We performed partial gastrectomy, and the resected specimen was diagnosed as needle tract seeding due to EUS-FNA. He underwent chemotherapy with TS-1, and he is still alive 18 months after surgery at the time of writing. CONCLUSION: For resectable pancreatic body or tail tumors, EUS-FNA should be carefully performed to prevent needle tract seeding and intraoperative as well as postoperative assessment for gastric wall metastasis is mandatory.

A prediction model for 5-year cardiac mortality in patients with chronic heart failure using ¹²³I-metaiodobenzylguanidine imaging.

PURPOSE: Prediction of mortality risk is important in the management of chronic heart failure (CHF). The aim of this study was to create a prediction model for 5-year cardiac death including assessment of cardiac sympathetic innervation using data from a multicenter cohort study in Japan. METHODS: The original pooled database consisted of cohort studies from six sites in Japan. A total of 933 CHF patients who underwent (123)I-metaiodobenzylguanidine (MIBG) imaging and whose 5-year outcomes were known were selected from this database. The late MIBG heart-to-mediastinum ratio (HMR) was used for quantification of cardiac uptake. Cox proportional hazard and logistic regression analyses were used to select appropriate variables for predicting 5-year cardiac mortality. The formula for predicting 5-year mortality was created using a logistic regression model. RESULTS: During the 5-year follow-up, 205 patients (22 %) died of a cardiac event including heart failure death, sudden cardiac death and fatal acute myocardial infarction (64 %, 30 % and 6 %, respectively). Multivariate logistic analysis selected four parameters, including New York Heart Association (NYHA) functional class, age, gender and left ventricular ejection fraction, without HMR (model 1) and five parameters with the addition of HMR (model 2). The net reclassification improvement analysis for all subjects was 13.8 % (p < 0.0001) by including HMR and its inclusion was most effective in the downward reclassification of low-risk patients. Nomograms for predicting 5-year cardiac mortality were created from the five-parameter regression model. CONCLUSION: Cardiac MIBG imaging had a significant additive value for predicting cardiac mortality. The prediction formula and nomograms can be used for risk stratifying in patients with CHF.

Indolent primary effusion lymphoma-like lymphoma in the pericardium: A case report and review of the literature.

We report a rare case of a 75-year-old man with a history of mild-to-moderate pericardial effusion that was detected on echocardiography performed in October 2011 when the patient was 69 years old. Follow-up echocardiography was performed every 6 months thereafter, showing that the pericardial effusion gradually subsided. However, in April 2017 he started experiencing several episodes of dyspnea, which prompted him to visit our hospital's outpatient department on June 22, 2017. Echocardiography revealed a large amount of pericardial effusion; thus, he was immediately hospitalized. After undergoing pericardiocentesis and drainage, 1740-ml of bloody pericardial fluid was collected. Serum antibody tests for human immunodeficiency virus, hepatitis C virus, and human herpes virus 8 were negative, whereas that for Epstein-Barr virus (EBV) was positive, indicating a prior infection. Cytopathological examination, immunocytochemical staining, lymphocyte surface marker analysis, and cytogenetic assessment were performed. EBV-encoded small ribonucleic acid in situ hybridization was negative. He was diagnosed with primary effusion lymphoma (PEL)-like lymphoma (LL) and was treated with 8 doses of rituximab 375 mg/m2 over a 2-month period. He has remained in complete response for the past 12 months. Our case shows the possibility of long-term existence of indolent PEL-LL in patients with mild-to-moderate pericardial effusion. .

Platelet Activation Assessed by Glycoprotein VI/Platelet Ratio Is Associated With Portal Vein Thrombosis After Hepatectomy and Splenectomy in Patients With Liver Cirrhosis.

Portal vein thrombosis (PVT) is a serious complication after hepatobiliary-pancreatic surgery. Portal vein thrombosis often develops in patients with liver cirrhosis (LC) postoperatively, although they have low platelet counts. Platelet activation is one of the causes of thrombosis formation, and soluble form of glycoprotein VI (sGPVI) has received attention as a platelet activation marker. We had prospectively enrolled the 81 consecutive patients who underwent splenectomy (Sx) and/or hepatectomy: these patients were divided as Sx (n = 38) and hepatectomy (Hx, n = 46) groups. The 3 patients who underwent both procedures were added to both groups. Each group was subdivided into patients with non-LC and LC: non-LC-Sx (n = 22) and LC-Sx (n = 16), non-LC-Hx (n = 40) and LC-Hx (n = 6). The presence of PVT was diagnosed by using enhanced computed tomography (CT) scan. Platelet counts were significantly lower in LC-Sx than in non-LC-Sx, and incidence of PVT was significantly higher in LC-Sx than in non-LC-Sx (68.8% vs 31.8%, P = .024). Soluble form of glycoprotein VI /platelet ratios on preoperative day and postoperative day 1 were significantly higher in LC-Sx than in non-LC-Sx. Incidence of PVT was significantly higher in LC-Hx than in non-LC-Hx (50.0% vs 7.5%, P < .01). Soluble form of glycoprotein VI /platelet ratios were significantly higher in LC-Hx before and after Hx, compared to non-LC-Hx. Patients with LC stay in hypercoagulable state together with platelet activation before and after surgery. Under this circumstance, alteration of portal venous blood flow after Sx or Hx is likely to cause PVT in patients with LC.

Cell patterning on polylactic acid through surface-tethered oligonucleotides.

Polylactic acid (PLA) is a candidate material to prepare scaffolds for 3-D tissue regeneration. However, cells do not adhere or proliferate well on the surface of PLA because it is hydrophobic. We report a simple and rapid method for inducing cell adhesion to PLA through DNA hybridization. Single-stranded DNA (ssDNA) conjugated to poly(ethylene glycol) (PEG) and to a terminal phospholipid (ssDNA-PEG-lipid) was used for cell surface modification. Through DNA hybridization, modified cells were able to attach to PLA surfaces modified with complementary sequence (ssDNA'). Different cell types can be attached to PLA fibers and films in a spatially controlled manner by using ssDNAs with different sequences. In addition, they proliferate well in a culture medium supplemented with fetal bovine serum. The coexisting modes of cell adhesion through DNA hybridization and natural cytoskeletal adhesion machinery revealed no serious effects on cell growth. The combination of a 3-D scaffold made of PLA and cell immobilization on the PLA scaffold through DNA hybridization will be useful for the preparation of 3-D tissue and organs.

A pooled analysis of multicenter cohort studies of (123)I-mIBG imaging of sympathetic innervation for assessment of long-term prognosis in heart failure.

OBJECTIVES: The study objectives were to create a cardiac metaiodobenzylguanidine (mIBG) database using multiple prospective cohort studies and to determine the quantitative iodine-123-labeled mIBG indices for identifying patients with chronic heart failure (HF) at greatest and lowest risk of lethal events. BACKGROUND: Although the prognostic value of cardiac mIBG imaging in patients with HF has been shown, clinical use of this procedure has been limited. It is required to define universally accepted quantitative thresholds for high and low risk that could be used as an aid to therapeutic decision-making using a large cohort database. METHODS: Six prospective HF cohort studies were updated, and the individual datasets were combined for the present patient-level analysis. The database consisted of 1,322 patients with HF followed up for a mean interval of 78 months. Heart-to-mediastinum ratio (HMR) and washout rate of cardiac mIBG activity were the primary cardiac innervation markers. The primary outcome analyzed was all-cause death. RESULTS: Lethal events were observed in 326 patients, and the population mortality rate was 5.6%, 11.3%, and 19.7% at 1, 2, and 5 years, respectively. Multivariate Cox proportional hazard model analysis for all-cause mortality identified age (p < 0.0001), New York Heart Association (NYHA) functional class (p < 0.0001), late HMR of cardiac mIBG activity (p < 0.0001), and left ventricular ejection fraction (LVEF) (p = 0.0029) as significant independent predictors. Analysis of the 512-patient subpopulation with B-type natriuretic peptide (BNP) results showed BNP (p < 0.0001), greater NYHA functional class (p = 0.0002), and late HMR (p = 0.0011) as significant predictors, but LVEF was not. The receiver-operating characteristic-determined threshold of HMR (1.68) identified patients at significantly increased risk in any LVEF category. Survival rates decreased progressively with decreasing HMR, with 5-year all-cause mortality rates >7% annually for HMR <1.25, and <2% annually for HMR ≥1.95. Addition of HMR to clinical information resulted in a significant net reclassification improvement of 0.175 (p < 0.0001). CONCLUSIONS: Pooled analyses of independent cohort studies confirmed the long-term prognostic value of cardiac mIBG uptake in patients with HF independently of other markers, such as NYHA functional class, BNP, and LVEF, and demonstrated that categoric assessments could be used to define meaningful thresholds for lethal event risk.

Prognostic value of repeated 123I-metaiodobenzylguanidine imaging in patients with dilated cardiomyopathy with congestive heart failure before and after optimized treatments--comparison with neurohumoral factors.

The present study was undertaken to assess whether repeated measurement of cardiac 123I-metaiodobenzylguanidine (MIBG) imaging parameters before and after optimized treatments is useful for predicting the prognosis of patients with congestive heart failure (CHF) resulting from dilated cardiomyopathy (DCM). The subjects were 85 consecutive patients with DCM who had a left ventricular ejection fraction (LVEF) of less than 45%. The MIBG and the concentrations of neurohumoral factors were measured at baseline and after 6 months of optimized treatments. Cox proportional hazards analysis was performed to assess the various parameters before and after treatment. Twenty-three patients had a cardiac event (12 died; 11 hospitalized) during a mean follow-up period of 2 years. Although there was no difference between the baseline heart to mediastinum (H/M) ratio measured by MIBG between survivors and nonsurvivors, the H/M ratio was significantly decreased in nonsurvivors after 6 months. Multivariate analysis revealed that a high plasma concentration of brain natriuretic peptide level after 6 months (p=0.0049) and absolute changes in the H/M ratio (p=0.0046) were independent predictors of mortality. Comparison of the H/M ratio on MIBG imaging before and after optimized additional treatment provided useful information for predicting mortality and was independent of clinical and neurohumoral factors previously shown to be associated with poor prognosis in patients with DCM.