Noncoding RNAs implication in cardiovascular diseases in the COVID-19 era.

COronaVIrus Disease 19 (COVID-19) is caused by the infection of the Severe Acute Respiratory Syndrome CoronaVirus 2 (SARS-CoV-2). Although the main clinical manifestations of COVID-19 are respiratory, many patients also display acute myocardial injury and chronic damage to the cardiovascular system. Understanding both direct and indirect damage caused to the heart and the vascular system by SARS-CoV-2 infection is necessary to identify optimal clinical care strategies. The homeostasis of the cardiovascular system requires a tight regulation of the gene expression, which is controlled by multiple types of RNA molecules, including RNA encoding proteins (messenger RNAs) (mRNAs) and those lacking protein-coding potential, the noncoding-RNAs. In the last few years, dysregulation of noncoding-RNAs has emerged as a crucial component in the pathophysiology of virtually all cardiovascular diseases. Here we will discuss the potential role of noncoding RNAs in COVID-19 disease mechanisms and their possible use as biomarkers of clinical use.

The hamstring muscle complex.

PURPOSE: The anatomical appearance of the hamstring muscle complex was studied to provide hypotheses for the hamstring injury pattern and to provide reference values of origin dimensions, muscle length, tendon length, musculotendinous junction (MTJ) length as well as width and length of a tendinous inscription in the semitendinosus muscle known as the raphe. METHODS: Fifty-six hamstring muscle groups were dissected in prone position from 29 human cadaveric specimens with a median age of 71.5 (range 45-98). RESULTS: Data pertaining to origin dimensions, muscle length, tendon length, MTJ length and length as well as width of the raphe were collected. Besides these data, we also encountered interesting findings that might lead to a better understanding of the hamstring injury pattern. These include overlapping proximal and distal tendons of both the long head of the biceps femoris muscle and the semimembranosus muscle (SM), a twist in the proximal SM tendon and a tendinous inscription (raphe) in the semitendinosus muscle present in 96 % of specimens. CONCLUSION: No obvious hypothesis can be provided purely based on either muscle length, tendon length or MTJ length. However, it is possible that overlapping proximal and distal tendons as well as muscle architecture leading to a resultant force not in line with the tendon predispose to muscle injury, whereas the presence of a raphe might plays a role in protecting the muscle against gross injury. Apart from these architectural characteristics that may contribute to a better understanding of the hamstring injury pattern, the provided reference values complement current knowledge on surgically relevant hamstring anatomy. LEVEL OF EVIDENCE: IV.

Postmortem imaging exposed: an aid in MR imaging of musculoskeletal structures.

OBJECTIVE: To identify factors that influence the quality of postmortem magnetic resonance (MR) images of musculoskeletal (MSK) structures as described in the literature, and to evaluate the extent to which these MR images are affected. MATERIALS AND METHODS: Four useful studies were retrieved from a PubMed and EMBASE search, covering the literature up to 1 March 2012. Three additional studies were included after a manual search from reference lists. RESULTS: Four human studies and three animal studies are considered in this review. Postmortem MRI quality can be affected by storage temperature, repeated freezing and thawing and fixation. Provided there was an adequate, but above-freezing storage temperature, postmortem changes in fresh cadavers did not appear to affect the MR image quality of MSK structures up to 14 days after death. Image contrast, signal intensities, and relaxation times are temperature-dependent, regardless of whether the specimen was fresh or postmortem for up to 7 days. Bad image quality can occur owing to accelerated autolysis. Freezing and thawing did not affect image quality, unless repeated too often, or whenever a heating pad was used to speed up the thawing process. Conventional formalin-based fixation leads to swelling of soft tissue and fluid accumulation in joints, and therefore to deteriorated images, with image quality just sufficient to visualize gross anatomy. CONCLUSION: Various factors were identified that affect postmortem MR image quality of MSK structures. Postmortem MR image quality was good, except for images of the fixated specimen. Freezing is the preferred method of conservation for specimens that are to be subjected to postmortem MRI.

Reconciling sectoral abatement strategies with global climate targets: the case of the Chinese passenger vehicle fleet.

The IPCC Forth Assessment Report postulates that global warming can be limited to 2 °C by deploying technologies that are currently available or expected to be commercialized in the coming decades. However, neither specific technological pathways nor internationally binding reduction targets for different sectors or countries have been established yet. Using the passenger car stock in China as example we compute direct CO(2) emissions until 2050 depending on population, car utilization, and fuel efficiency and compare them to benchmarks derived by assuming even contribution of all sectors and a unitary global per capita emission quota. Compared to present car utilization in industrialized countries, massive deployment of prototypes of fuel efficient cars could reduce emissions by about 45%, and moderately lower car use could contribute with another 33%. Still, emissions remain about five times higher than the benchmark for the 2 °C global warming target. Therefore an extended analysis, including in particular low-carbon fuels and the impact of urban and transport planning on annual distance traveled and car ownership, should be considered. A cross-sectoral comparison could reveal whether other sectors could bear an overproportional reduction quota instead. The proposed model offers direct interfaces to material industries, fuel production, and scrap vehicle supply.

E17K substitution in AKT1 in prostate cancer.

BACKGROUND: The phosphatidylinositol 3-kinase (PI3K)-AKT pathway is activated in many cancers. Mutational hotspots in AKT1 and in the regulatory and catalytic subunits of PI3K have been detected in multiple tumour types. In AKT1, the E17K substitution leads to a PI3K-independent activation of AKT1. METHODS: A mutational profiling of AKT1 and of the mutational hotspots in PIK3CA and PIK3R1 was carried out in samples from primary and recurrent prostate tumours. RESULTS: We show that, in prostate cancer, AKT1(E17K) had a prevalence of 1.4%. The mutation seemed to be associated with a favourable clinical course but it was not associated with a specific tumour growth pattern. Activating mutations in PIK3CA or PIK3R1 were not found in prostate cancer. CONCLUSION: The E17K substitution in AKT1 is rare in prostate cancer. It seems associated with a favourable clinical outcome but not with a specific histology of the tumour.

PTEN-mediated G1 cell-cycle arrest in LNCaP prostate cancer cells is associated with altered expression of cell-cycle regulators.

BACKGROUND: The tumor suppressor PTEN regulates many biological processes. A well-known downstream effector of PTEN is phospho-Akt. Although PTEN is the most frequently inactivated gene in prostate cancer, its mode of action is not fully understood. We studied the association of regulated PTEN expression with changes in biological function and gene expression profiles. METHODS: PTEN-negative LNCaP cells were stably transfected with wild-type PTEN cDNA under inducible control, resulting in LNCaP/PTEN cells. Microarray analysis was used to monitor gene expression changes upon induction of PTEN. Expression of selected individual genes was studied in Q-PCR and siRNA experiments. Cell-cycle distribution was analyzed by flow cytometry. RESULTS: Induced expression of PTEN in LNCaP/PTEN cells significantly inhibited cell proliferation, at least partly due to cell-cycle arrest at the G1 phase. Expression profiling combined with pathway analysis revealed that PTEN-dependent G1 growth arrest was associated with an altered mRNA expression of the G1 cell-cycle regulators Cdc25a, E2F2, cyclin G2, and RBL2/p130. Specific inhibition of Akt signaling by siRNA resulted in downregulation of both E2F2 and Cdc25a mRNA expression and upregulation of the FOXO target cyclin G2, similar to the effect observed by PTEN induction. However, Akt did not mediate the PTEN-dependent RBL2/p130 mRNA expression in LNCaP/PTEN cells. CONCLUSIONS: The results indicate that PTEN dependent gene expression is important in cell-cycle regulation and is mediated by both Akt-dependent and -independent mechanisms.

Commutability of control materials in glycohemoglobin determinations.

The intermethod variabilities of control materials and patient blood samples for the measurement of glycohemoglobin were compared. Sets of 50 blood samples and 15 control materials were analyzed by HPLC and affinity and immunochemical methods. For each pair of methods, the distances of the materials from the regression line of patient blood results (expressed as normalized residuals) were calculated. Only two of 15 controls had normalized residuals exceeding 3 standard deviations from the regression line. Total hemoglobin (Hb) content, Hb derivatives, and cellulose acetate electrophoresis demonstrated that only a minority of controls could be considered similar to patients' blood samples. We selected Menarini's and our home-prepared controls to simulate calibration of the different techniques by these materials. Intermethod calibration succeeded mostly in harmonizing results obtained by HPLC methods. On the contrary, calibration of the immunochemical techniques (Boehringer and Roche) did not improve intermethod agreement to a clinically useful level.

A novel gene which is up-regulated during colon epithelial cell differentiation and down-regulated in colorectal neoplasms.

To identify new molecular markers for differentiation of normal and neoplastic colon epithelium, we have studied changes in gene expression during the in vitro differentiation of the HT29-D4 colon carcinoma cell line. Using a modified differential display procedure, we cloned a novel cDNA, designated differentiation-related gene 1 (Drg1). Drg1 mRNA has a length of approximately 3 kb and is induced approximately 20-fold during in vitro differentiation of the colon carcinoma cell lines HT29-D4 and Caco-2. The absence of Drg1 induction in growth-inhibited A431 epidermoid carcinoma cells indicates that Drg1 up-regulation in colon carcinoma cells is not a result of decreased proliferation. The Drg1 cDNA contains an open-reading frame of 1182 bp that encodes a protein with a predicted molecular weight of 43 kd. Drg1 mRNA is expressed most prominently in placental membranes and prostate, kidney, small intestine, and ovary tissues. Compared to normal colon mucosa, Drg1 mRNA expression is decreased in colon adenomas and adenocarcinomas. An antiserum raised against recombinant Drg1 protein detected a band of the expected size in Western blots. Immunohistochemistry showed that in normal colon Drg1 protein is expressed in the cytoplasm and basolateral membranes of surface epithelial cells that border the gut lumen, indicating that Drg1 protein is expressed late during differentiation, just before apoptosis and shedding of cells into the colon lumen.

Androgen regulation of the rat keratinocyte growth factor (KGF/FGF7) promoter.

Keratinocyte Growth Factor (KGF/FGF7) is a candidate andromedin in normal embryonic development of male accessory sex glands, such as the prostate and seminal vesicles. The expression of KGF mRNA and protein is androgen-responsive. To elucidate the regulation of expression of the KGF gene, we isolated the first two exons of the KGF gene and approximately 15 kb upstream sequences. The major transcription start site was mapped. It is preceded by a CAAT-box and a TATA-box. Transient transfections in LNCaP cells revealed that, upon treatment with the synthetic androgen R1881, KGF promoter activity is upregulated 6 to 11 fold, indicating androgen regulation of the KGF promoter in the region from position - 900 to -1200. The longest construct (BH-pLuc: -4700 to +901) has a much higher basal activity than the shorter constructs, indicating that in the region -4700 to -2700 additional activating sequences are present.

Expression of nucleophosmin/B23 in normal and neoplastic colorectal mucosa.

Nucleophosmin/B23 is a 38 kD molecular phosphoprotein involved in ribosome assembly and transport. In view of the fact that nucleophosmin/B23 appears to be more abundant in tumour cells than in normal cells, the mRNA expression and immunohistochemical localization of nucleophosmin/B23 were investigated in 19 samples of non-neoplastic mucosa, six adenomas, and 16 adenocarcinomas of the colorectum. Northern blot analysis revealed that nucleophosmin/B23 mRNA is expressed at a higher level in adenomas and carcinomas than in non-neoplastic mucosa of the colorectum. Immunohistochemical staining of formalin-fixed, paraffin-embedded tissue sections after microwave antigen retrieval, using a nucleophosmin/B23-specific monoclonal antibody, showed almost exclusively diffuse nuclear reactivity of a majority of the epithelial cells in non-neoplastic mucosa: in adenomas, reactivity was almost exclusively nucleolar and in carcinomas, nuclear as well as nucleolar staining was observed. During mitosis, the immunoreactivity of nucleophosmin/B23 appears in the cytoplasm. The results indicate that the expression of nucleophosmin/B23 is higher in neoplastic than in non-neoplastic colorectal mucosa. Furthermore, the pattern of nucleophosmin/B23 expression shifts from nuclear to nucleolar early in the adenoma-carcinoma sequence. The exact function of nucleophosmin/B23 in colorectal carcinogenesis remains to be determined.

Identification of mRNAs that show modulated expression during colon carcinoma cell differentiation.

To investigate the working hypotheses that stem cells or their early descendants are prime targets for neoplastic transformation, and that the degree to which a neoplasm retains the immature phenotype is an important determinant of tumor aggressiveness, we have identified several mRNAs that are downregulated during the in vitro differentiation of HT29-D4 colon carcinoma cells. These genes include heat-shock cognate protein Hsc70, adenylosuccinate lyase, B23/nucleophosmin, alpha-tubulin, and a novel gene designated DS-1. The DS-1 mRNA has a length of approximately 0.9 kb and is downregulated 4.7-fold upon differentiation. From the DS-1 cDNA, a protein of 206 amino acids with a molecular mass of 24 kDa and an isoelectric point of 10.9 can be deduced. An antiserum directed against a synthetic peptide detected a minor band of the expected size in Western blots, as well as a major band of lower size that may represent a processed form of the protein.

Combinatorial association and abundance of components of interferon-stimulated gene factor 3 dictate the selectivity of interferon responses.

Genes containing the interferon-stimulated response element (ISRE) enhancer have been characterized as transcriptionally responsive primarily to type I interferons (IFN alpha/beta). Induction is due to activation of a multimeric transcription factor, interferon-stimulated gene factor 3 (ISGF3), which is activated by IFN alpha/beta but not by IFN gamma. We found that ISRE-containing genes were induced by IFN gamma as well as by IFN alpha in Vero cells. The IFN gamma response was dependent on the ISRE and was accentuated by preexposure of cells to IFN alpha, a treatment that increases the abundance of ISGF3 components. Overexpression of ISGF3 polypeptides showed that the IFN gamma response depended on the DNA-binding protein ISGF3 gamma (p48) as well as on the 91-kDa protein STAT91 (Stat1 alpha). The transcriptional response to IFN alpha required the 113-kDa protein STAT113 (Stat2) in addition to STAT91 and p48. Mutant fibrosarcoma cells deficient in each component of ISGF3 were used to confirm that IFN gamma induction of an ISRE reporter required p48 and STAT91, but not STAT113. A complex containing p48 and phosphorylated STAT91 but lacking STAT113 bound the ISRE in vitro. IFN gamma-induced activation of this complex, preferentially formed at high concentrations of p48 and STAT91, may explain some of the overlapping responses to IFN alpha and IFN gamma.

The interferon-stimulated gene 54 K promoter contains two adjacent functional interferon-stimulated response elements of different strength, which act synergistically for maximal interferon-alpha inducibility.

The interferon-alpha(IFN-alpha)-regulated hamster ISG-54 K gene, which is activated in hamster CHO-12 cells at least 40-fold, was isolated and the promoter region was characterized in detail. Sequence analysis revealed the presence of two elements, closely related to the interferon-stimulated-response-element (ISRE) consensus sequence [AGTTTCNNTTTC(CT)]. The putative ISRE-I sequence (GGTTTCAATTTCT) is located at position -97 to -85; ISRE-II (AGTTTTACTTTCT), which differs at three positions from ISRE-I, is found directly upstream of ISRE-I at position -110 to -98. In a transient transfection assay in CHO-12 cells the wild-type hamster ISG-54K-promoter-chloramphenicol-acetyl-transferase (CAT) reporter construct showed a 40-80-fold induction, offering an excellent model to study the functional properties of the two ISRE. To find out whether both elements were functional in interferon regulation of the promoter, selected point mutations were introduced in the -110 to -85 region and in flanking sequences. The (mutated) ISG-54 K promoter was linked to the CAT reporter gene and transiently expressed in CHO cells in the absence and presence of murine (Mu)IFN-alpha 6. Transfections showed that both the -97 to -85 (ISRE-I) and the -110 to -98 (ISRE-II) segment were needed for optimal interferon induction of the ISG-54 K promoter. However, ISRE-I has an approximately sevenfold stronger activity compared to ISRE-II. Sequential substitution of the three ISRE-I bases, which differ in ISRE-II showed that the T at position -105 causes the lower activity of ISRE-II. Transfection of ISG-54 K promoter constructs, in which ISRE-I was replaced by ISRE-II, which generates a promoter with two ISRE-II segments, and vice versa (two ISRE-I), provided further evidence for a role of both elements in IFN-alpha induction. Importantly, all data obtained in transfection studies show that the two ISRE cooperate synergistically. The mechanism of synergism is most probably an indirect interaction between transcription factors binding to the ISRE, because an increase in the spacial arrangement of the two ISRE with a complete helical turn or half a turn did not result in a substantial decrease in promoter activity.

[Perforated duodenal ulcer in children]. / Ulcus duodenal perforado en la infancia.

We face a case of duodenal ulcer perforation a nine year old child who's diagnosis was not detected before operating. This is an unusual case and it is not normally observed in the differential diagnosis of the acute abdominal pain in the infancy. The lack of the radiological signs of perforation contributed to the diagnosis mistake before operating.