Probes for INS

Aberrant Ca-CaM signaling has been implicated in various congenital and acquired cardiac pathologies, including arrhythmia, hypertrophy, and HF. We examined the impact of HF induced by trans-aortic constriction (TAC) on the distribution of the three CaM mRNAs (Calm 1,2 and 3) and their key protein target mRNAs (Ryr2, Scn5a, Camk2d, NOS1 and Cacna1c) in cardiomyocytes, using fluorescence in situ hybridization (RNAScope™). HF resulted in specific changes in the pattern of localization of Calms, manifested in redistribution of Calm3 from the cell periphery towards the perinuclear area and enhanced Calm2 attraction to the perinuclear area compared to sham myocytes. Additionally, HF resulted in redistribution of mRNAs for certain CaM target mRNAs. Particularly, NOS1 localization shifted from the cell periphery towards the perinuclear area, Cacna1c, Camk2d and Scn5a abundance increased at the perinuclear area, and Ryr2 attracted even closer to the cell periphery in HF myocytes compared to sham myocytes. The strength of non-random attraction/repulsion was measured as the maximal deviation between the observed distribution of nearest neighbor distances from the distribution predicted under complete spatial randomness. Consistent with the observed alterations in abundance and distribution of CaM and CaM target mRNAs, HF resulted in increased attraction between Calm1 and Scn5a, Ryr2 and Camk2d, between Calm2 and Ryr2 and Camk2d; and between Calm3 and NOS1 and Scn5a. In contrast, the attraction between Calm3 and Ryr2 decreased in HF myocytes compared to sham. Collectively, these results suggest distribution of Calms and their association with key target protein mRNAs undergo substantial alterations in heart failure. These results have new important implications for organization of Ca signaling in normal and diseased heart.

Objective: To assess the clinical features and treatment outcomes in patients with primary ovarian squamous cell carcinoma (POSCC). Methods: Fifteen patients with primary ovarian squamous cell carcinoma diagnosed from January 2009 to December 2018 in Cancer Hospital of the University of Chinese Academy of Sciences were collected. The expression of p16, hMLH1, hMSH2, hMSH6 and PMS2 in POSCC was detected by immunohistochemistry, and the status of high-risk human papillomavirus (HPV) by RNAscope test. Results: Squamous cell carcinoma with different degrees of differentiation was found in 15 cases, including three cases with high differentiation and 12 cases with medium to low differentiation. There were four cases with in situ squamous cell carcinoma, four cases with teratoma, one case with endometrial carcinoma/atypical hyperplasia, and one case with endometriosis. p16 was expressed in five cases (5/15), indicating coexisting high-risk HPV infection. There was no high-risk HPV infection in the remaining 10 cases, and p16 staining was negative. There was no deficient mismatch repair protein in all cases. The overall survival time (P=0.038) and progression free survival (P=0.045) of patients with high-risk HPV infection were longer than those without HPV infection. Conclusions: POSCC is more commonly noted in postmenopausal women and often occurs unilaterally. Elevated serological indexes CA125 and SCC are the most common finding. Morphologically, the tumors show variable degrees of differentiation, but the current data suggest that the degree of differentiation cannot be used as an independent prognostic index. High-risk HPV infection may be associated with the occurrence of POSCC, and that the prognosis of POSCC patients with HPV infection is better than that of patients without infection.

Atrial fibrillation (AF) is commonly observed in patients with hypertension and is associated with pathologically elevated cardiomyocyte stretch. AF triggers have been linked to subcellular Ca2+ abnormalities, while their association with stretch remains elusive. Caveolae are mechanosensitive membrane structures, that play a role in both Ca2+ and cyclic adenosine monophosphate (cAMP) signaling. Therefore, caveolae could provide a mechanistic connection between cardiomyocyte stretch, Ca2+ mishandling, and AF. In isolated mouse atrial myocytes, cell stretch was mimicked by hypotonic swelling, which increased cell width (by ∼30%, p

Gastric-type cervical adenocarcinoma (GCA) is a rare and aggressive type of endocervical adenocarcinoma (ECA) with distinct histopathologic features and unfavorable treatment outcomes, but no genomic prognostic factor has been revealed. We aimed to systematically investigate the somatic alterations of GCA at genome-wide level and evaluate their prognostic value.We performed whole-exome sequencing (WES) on 25 pairs of tumor and matched normal samples to characterize the genomic features of Chinese patients with GCA and investigated their relations to histopathological characterizations and prognosis. The prognostic value of the genomic alterations was evaluated in a total of 58 GCA patients.Mutations were commonly observed in reported GCA-related driver genes, including TP53 (32%), CDKN2A (20%), SKT11 (20%), BRCA2 (12%), SMAD4 (12%), and ERBB2 (12%). Recurrent novel trunk mutations were also observed in PBRM1 (12%), FRMPD4 (12%), and NOP2 (8%) with high variant allele frequency. Moreover, enrichment of the APOBEC signature was attributed to frequent gain of somatic copy number alteration (SCNA) of APOBEC3B (20%), which perfectly matched the nuclear-positive staining of APOBEC3B through immunohistochemistry. In contrast, APOBEC3B alteration was absent in patients with conventional type of ECA (N = 52). Notably, positive APOBEC3B was consistently enriched in patients with favorable prognosis in both the discovery cohort and an additional 33 GCA patients, thus indicating a significant association with lower relapse risk of GCA independent of cancer stage (P = 0.02).Our results can aid understanding of the molecular basis of GCA in the Chinese population by providing genomic profiles and highlighting the potential prognostic value of APOBEC3B for GCA through routine clinical IHC.

Stochastic spiking is a prominent feature of Ca2+ signaling. The main noise source at the cellular level are puffs from inositol-trisphosphate receptor (IP3R) channel clusters in the membrane of the endoplasmic reticulum (ER). While the random cluster activity has been known for decades, a stringent method to derive the puff noise term acting on the cytosolic Ca2+ concentration is still lacking. We adopt a popular description of neural spike generation from neuroscience, the stochastic integrate-and-fire (IF) model, to describe Ca2+ spiking. Our model consists of two components describing i) activity of IP3R clusters and ii) dynamics of the global Ca2+ concentrations in the cytosol and in the ER. Cluster activity is modeled by a Markov chain, capturing the puff. The global Ca2+ concentrations are described by a two-variable IF model driven by the puff current. For the Markov chain we derive expressions for the statistics of interpuff interval, single-puff strength, and puff current assuming constant cytosolic Ca2+, an assumption often well met because the Ca2+ concentrations vary much slower than the cluster activity does. The latter assumption also allows to approximate the driving Ca2+ dependent puff current by a white Gaussian noise. This approximation results in an IF model with nonlinear drift and multiplicative noise. We consider this reduced model in a renewal version and in a version with cumulative refractoriness. Neglecting ER depletion, the stochastic IF model has only one variable and generates a renewal spike train, a point process with statistically independent interspike intervals (ISI). We derive analytical expressions for the mean and coefficient of variation of the ISI and suggest approximations for the ISI density and spike-train power spectrum. Taking into account ER depletion, the two-variable IF model displays cumulative refractoriness as seen in experimental data.

Independent primary uterine and cervical adenocarcinoma are rare and difficult to identify their origins, which makes treatment decision difficult. A 46-year-old female with endometrioid carcinoma and adenocarcinoma, human papilloma virus (HPV)-associated of the uterine cervix was reported. The patient presented with increased menstrual flow, contact bleeding and watery leucorrhea for more than one year, and the imaging findings showed abnormal uterine morphology, irregular margins, and multiple abnormal signals in uterine cavity and myometrium, which suggested multiple leiomyomas of the uterus. The signal intensity in the right muscle layer was markedly enhanced, suggesting a smooth muscle tumor of uncertain malignant potential. A large number of cystic hypointensity was seen in the cervix, and multiple cysts were considered. The initial preoperative diagnosis was multiple leiomyoma of the uterus, and a hysterectomy operation was planned. During the operation, the uterus was sent for frozen sections. There was a mass in the endometrium of the fundus, with a soft grayish-red cut surface and a clear border with the myometrium, and there was a grayish-white nodule in the cervix with a hard grayish-white cut surface. The two masses were well demarcated from each other, and the distance between them was 30 mm. The result of the frozen sections indicated the malignant tumor of the endometrium, and the extended hysterectomy+pelvic lymphadenectomy+partial resection of the greater omentum was performed. After the operation, the paraffin sections were sent to the Department of Pathology of the Peking University Third Hospital for histochemistry, POLE gene sequencing and HPV RNAscope tests, and the final diagnosis was a synchronous endometrioid carcinoma (POLE-mutant according to the WHO classification) and an adenocarcinoma, HPV-associated of the uterine cervix. Now the patient had been treated with 2 cycles of chemotherapy and her condition was fine. Through the analysis of the histological, immunohistochemical and molecular detection results of this case, the importance of applying HPV RNAscope and TCGA molecular typing in the diagnosis of cervical adenocarcinomas and endometrial carcinomas was emphasized. At the same time, gynecologists should not blindly rely on intraoperative frozen sections, and should pay attention to preoperative pathological examination, and make appropriate operation methods according to the results in order to prevent passivity in the surgery.

Primary female urethral carcinoma (PUC-F) accounts for less than 1% of all genitourinary malignancies and comprises a histologically diverse group of tumors that are usually associated with poor prognosis. The carcinomas documented at this site include adenocarcinoma (clear cell adenocarcinoma, columnar cell carcinoma, and Skene gland adenocarcinoma), urothelial carcinoma (UCa), and squamous cell carcinoma (SCC). Recent studies have shown adenocarcinomas to be the most common type of primary urethral carcinoma in females. As most of the urethral carcinomas morphologically resemble carcinomas arising from surrounding pelvic organs or metastases, these should be ruled out before making the diagnosis of PUC-F. These tumors are currently staged according to the 8th edition of the American Joint Committee on Cancer (AJCC) staging system. However, the AJCC system has limitations, including the staging of tumors involving the anterior wall of the urethra. Staging systems like the recently proposed histology-based female urethral carcinoma staging system (UCS) takes into account the unique histological landmarks of the female urethra to better stratify pT2 and pT3 tumors into prognostic groups, that correlate with clinical outcomes including recurrence rates, disease-specific survival and overall survival. Further larger multi-institutional cohorts are however required to validate the results of this staging system. There is very limited information regarding the molecular profiling of PUC-F. Thirty-one percent of clear cell adenocarcinomas have been reported to show PIK3CA alterations, whereas 15% of adenocarcinomas show PTEN mutations. Higher tumor mutational burden and PD-L1 staining have been reported in UCa and SCC. Although multimodality treatment is usually recommended in locally advanced and metastatic disease, the role of immunotherapy and targeted therapy is promising in select PUC-F cases.

The mitochondrial calcium uniporter is a multi-subunit calcium channel that imports Ca2+ into mitochondria. Its MICU subunits (MICU1, MICU2, and the neuron-specific MICU3) gate the channel by blocking the pore in low Ca2+. Upon local Ca2+ elevation, Ca2+ binds to MICUs to cause MICU unblock, thus opening the pore so Ca2+ can permeate. Previous work using cell lines suggests that the uniporter in mammalian cells is exclusively regulated by a MICU1-MICU2 heterodimer. However, we show here that multiple types of electrically excitable cells, including skeletal muscle and cardiac tissues, can also possess a MICU1-MICU1 homodimer or virtually no MICUs. Kinetic analyses demonstrate that MICU1 has a higher Ca2+ affinity than MICU2, and that without MICUs the uniporter is constitutively open. As a result, uniporters with the MICU1-1 homodimer or no MICUs exhibit higher transport activities, leading to mitochondria accumulating much higher levels of matrix Ca2+. Using a Seahorse assay, we show that cells with MICU1-1 or no MICUs have impaired basal oxidative phosphorylation, likely due to increased ROS and damaged respiratory-complex proteins, including NDUFS3 and COX2. These cells, moreover, are highly susceptible to apoptosis. The disadvantage of employing MICU1-1 or omitting MICUs, however, accompanies strong physiological benefits. We show that in response to intracellular Ca2+ signals, these mitochondria import more Ca2+ and consequently produce more ATP, thus better supplying the energy required for the cellular processes initiated by the Ca2+ signals. In conclusion, this work reveals that tissues can manipulate their mitochondrial calcium uptake properties to suit their unique physiological needs by customizing their MICU regulation of the mitochondrial calcium uniporter.

The incorporation of HPV DNA testing into cervical screening programs has shown that many HPV-positive women are cytologically normal, with HPV-positivity fluctuating throughout life. Such results suggest that papillomaviruses may persist in a latent state after disease clearance, with sporadic recurrence. It appears that virus latency represents a narrow slot in a wider spectrum of subclinical and possibly productive infections. Clinical studies, and animal model infection studies, suggested a key role for host immune surveillance in maintaining such asymptomatic infections, and although infections may also be cleared, most studies have used the term 'clearance' to describe a situation where the presence of HPV DNA falls below the clinical detection level. Given our knowledge of papillomavirus immune evasion strategies and the restricted pattern of viral gene expression required for 'basal cell' persistence, the term 'apparent clearance' and 'subclinical persistence' of infection may better summarise our understanding. Subclinical infection also encompasses the lag phase, which occurs between infection and lesion development. This is dependent on infection titre, with multifocal infections developing more rapidly to disease. These concepts can usefully influence patient management where HPV-positivity occurs sometime after the onset of sexual activity, and where vertical transmission is suspected despite a lag period.