Plasma membrane injury can cause lethal influx of calcium, but cells survive by mounting a polarized repair response targeted to the wound site. Mitochondrial signaling within seconds after injury ...enables this response. However, as mitochondria are distributed throughout the cell in an interconnected network, it is unclear how they generate a spatially restricted signal to repair the plasma membrane wound. Here we show that calcium influx and Drp1-mediated, rapid mitochondrial fission at the injury site help polarize the repair response. Fission of injury-proximal mitochondria allows for greater amplitude and duration of calcium increase in these mitochondria, allowing them to generate local redox signaling required for plasma membrane repair. Drp1 knockout cells and patient cells lacking the Drp1 adaptor protein MiD49 fail to undergo injury-triggered mitochondrial fission, preventing polarized mitochondrial calcium increase and plasma membrane repair. Although mitochondrial fission is considered to be an indicator of cell damage and death, our findings identify that mitochondrial fission generates localized signaling required for cell survival.
Alternative pre-messenger RNA splicing influences development, physiology and disease, but its regulation in humans is not well understood, partially because of the limited scale at which the ...expression of specific splicing events has been measured. We generated the first genome-scale expression compendium of human alternative splicing events using custom whole-transcript microarrays monitoring expression of 24,426 alternative splicing events in 48 diverse human samples. Over 11,700 genes and 9,500 splicing events were differentially expressed, providing a rich resource for studying splicing regulation. An unbiased, systematic screen of 21,760 4-mer to 7-mer words for cis-regulatory motifs identified 143 RNA 'words' enriched near regulated cassette exons, including six clusters of motifs represented by UCUCU, UGCAUG, UGCU, UGUGU, UUUU and AGGG, which map to trans-acting regulators PTB, Fox, Muscleblind, CELF/CUG-BP, TIA-1 and hnRNP F/H, respectively. Each cluster showed a distinct pattern of genomic location and tissue specificity. For example, UCUCU occurs 110 to 35 nucleotides preceding cassette exons upregulated in brain and striated muscle but depleted in other tissues. UCUCU and UGCAUG seem to have similar function but independent action, occurring 5' and 3', respectively, of 33% of the cassette exons upregulated in skeletal muscle but co-occurring for only 2%.
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Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
The rapidly advancing field of digital health technologies provides a great opportunity to radically transform the way clinical trials are conducted and to shift the clinical trial paradigm from a ...site‐centric to a patient‐centric model. Merck’s (Kenilworth, NJ) digitally enabled clinical trial initiative is focused on introduction of digital technologies into the clinical trial paradigm to reduce patient burden, improve drug adherence, provide a means of more closely engaging with the patient, and enable higher quality, faster, and more frequent data collection. This paper will describe the following four key areas of focus from Merck’s digitally enabled clinical trials initiative, along with corresponding enabling technologies: (i) use of technologies that can monitor and improve drug adherence (smart dosing), (ii) collection of pharmacokinetic (PK), pharmacodynamic (PD), and biomarker samples in an outpatient setting (patient‐centric sampling), (iii) use of digital devices to collect and measure physiological and behavioral data (digital biomarkers), and (iv) use of data platforms that integrate digital data streams, visualize data in real‐time, and provide a means of greater patient engagement during the trial (digital platform). Furthermore, this paper will discuss the synergistic power in implementation of these approaches jointly within a trial to enable better understanding of adherence, safety, efficacy, PK, PD, and corresponding exposure‐response relationships of investigational therapies as well as reduced patient burden for clinical trial participation. Obstacle and challenges to adoption and full realization of the vision of patient‐centric, digitally enabled trials will also be discussed.
We developed a procedure for the preparation of whole transcriptome cDNA libraries depleted of ribosomal RNA from only 1 μg of total RNA. The method relies on a collection of short, computationally ...selected oligonucleotides, called 'not-so-random' (NSR) primers, to obtain full-length, strand-specific representation of nonribosomal RNA transcripts. In this study we validated the technique by profiling human whole brain and universal human reference RNA using ultra-high-throughput sequencing.
Merck & Co, Inc (Kenilworth, NJ) is investing in approaches to enrich clinical trial data and augment decision making through use of digital health technologies, outpatient sampling, and real‐time ...data access. As part of this strategy, a phase I study was conducted to explore a few technologies of interest. In this fixed‐sequence two‐period trial, 16 healthy subjects were administered 50‐mg once‐daily sitagliptin packaged in a bottle that electronically captured the date and time study medication was dispensed (period 1) and in a traditional pharmacy bottle (period 2). Dried blood spot samples were collected for sitagliptin concentration analysis on select study days, both in clinic and at home, with collection time recorded using an electronic diary in period 1 and by clinic staff in period 2. Study results demonstrated the feasibility and subject acceptance of collecting digital adherence data and outpatient dried blood spot samples in clinical trials and highlighted areas for future improvements.
After diagnosis of a cardiac mass, clinicians must weigh the benefits and risks of ascertaining a tissue diagnosis. Limited data are available on the accuracy of previously developed noninvasive ...pediatric cardiac magnetic resonance (CMR)-based diagnostic criteria.BACKGROUNDAfter diagnosis of a cardiac mass, clinicians must weigh the benefits and risks of ascertaining a tissue diagnosis. Limited data are available on the accuracy of previously developed noninvasive pediatric cardiac magnetic resonance (CMR)-based diagnostic criteria.The goals of this study were to: 1) evaluate the CMR characteristics of pediatric cardiac masses from a large international cohort; 2) test the accuracy of previously developed CMR-based diagnostic criteria; and 3) expand diagnostic criteria using new information.OBJECTIVESThe goals of this study were to: 1) evaluate the CMR characteristics of pediatric cardiac masses from a large international cohort; 2) test the accuracy of previously developed CMR-based diagnostic criteria; and 3) expand diagnostic criteria using new information.CMR studies (children 0-18 years of age) with confirmatory histological and/or genetic diagnosis were analyzed by 2 reviewers, without knowledge of prior diagnosis. Diagnostic accuracy was graded as: 1) single correct diagnosis; 2) correct diagnosis among a differential; or 3) incorrect diagnosis.METHODSCMR studies (children 0-18 years of age) with confirmatory histological and/or genetic diagnosis were analyzed by 2 reviewers, without knowledge of prior diagnosis. Diagnostic accuracy was graded as: 1) single correct diagnosis; 2) correct diagnosis among a differential; or 3) incorrect diagnosis.Of 213 cases, 174 (82%) had diagnoses that were represented in the previously published diagnostic criteria. In 70% of 174 cases, both reviewers achieved a single correct diagnosis (94% of fibromas, 71% of rhabdomyomas, and 50% of myxomas). When ≤2 differential diagnoses were included, both reviewers reached a correct diagnosis in 86% of cases. Of 29 malignant tumors, both reviewers indicated malignancy as a single diagnosis in 52% of cases. Including ≤2 differential diagnoses, both reviewers indicated malignancy in 83% of cases. Of 6 CMR sequences examined, acquisition of first-pass perfusion and late gadolinium enhancement were independently associated with a higher likelihood of a single correct diagnosis.RESULTSOf 213 cases, 174 (82%) had diagnoses that were represented in the previously published diagnostic criteria. In 70% of 174 cases, both reviewers achieved a single correct diagnosis (94% of fibromas, 71% of rhabdomyomas, and 50% of myxomas). When ≤2 differential diagnoses were included, both reviewers reached a correct diagnosis in 86% of cases. Of 29 malignant tumors, both reviewers indicated malignancy as a single diagnosis in 52% of cases. Including ≤2 differential diagnoses, both reviewers indicated malignancy in 83% of cases. Of 6 CMR sequences examined, acquisition of first-pass perfusion and late gadolinium enhancement were independently associated with a higher likelihood of a single correct diagnosis.CMR of cardiac masses in children leads to an accurate diagnosis in most cases. A comprehensive imaging protocol is associated with higher diagnostic accuracy.CONCLUSIONSCMR of cardiac masses in children leads to an accurate diagnosis in most cases. A comprehensive imaging protocol is associated with higher diagnostic accuracy.
Abstract
Background
Primary cystic neoplasms of the peritoneum are rare lesions and not commonly encountered in practice. Many intra-abdominal processes may mimic cystic masses within the peritoneal ...cavity and pose a diagnostic challenge to both the pathologist and radiologist. Clinical presentation is diverse and varied. These lesions are usually benign. Hence complete surgical excision is the treatment of choice in most of the cases.
Methods
Study design: Descriptive Retrospective study.
Cystic peritoneal lesions were identified and studied from data over a period of 5 years in the Histopathology Section at a tertiary care hospital in Pune, India. Mode of presentation, imaging findings in addition to gross and histopathologic findings of these lesions were studied.
Results
Out of 50 peritoneal lesions studied over a period of 5 years, only 7 were identified to be cystic peritoneal masses.
Of these two were found to be peritoneal cysts, two mesenteric cysts, one an infected mesenteric cyst and one each a mucinous cystadenoma and lymphangioma.
Conclusions
Correct diagnosis rests in the hands of the pathologist and ensures that the patient receives appropriate and timely management. Hence knowledge of the spectrum of these rare cystic peritoneal masses is necessary to distinguish from other potential cystic abdominal mimicker masses and avoid a potential pitfall.
RNA interference has revolutionized our ability to study the effects of altering the expression of single genes in mammalian (and other) cells through targeted knockdown of gene expression. In this ...report we describe a web-based computational tool, siRNA Information Resource (sIR), which consists of a new open source database that contains validation information about published siRNA sequences and also provides a user-friendly interface to design and analyze siRNA sequences against a chosen target sequence.
The siRNA design tool described in this paper employs empirically determined rules derived from a meta-analysis of the published data; it uses a weighted scoring system that determines the optimal sequence within a target mRNA and thus aids in the rational selection of siRNA sequences. This scoring system shows a non-linear correlation with the knockdown efficiency of siRNAs. sIR provides a fast, customized BLAST output for all selected siRNA sequences against a variety of databases so that the user can verify the uniqueness of the design. We have pre-designed siRNAs for all the known human genes (24,502) in the Refseq database. These siRNAs were pre-BLASTed against the human Unigene database to estimate the target specificity and all results are available online.
Although most of the rules for this scoring system were influenced by previously published rules, the weighted scoring system provides better flexibility in designing an appropriate siRNA when compared to the un-weighted scoring system. sIR is not only a comprehensive tool used to design siRNA sequences and lookup pre-designed siRNAs, but it is also a platform where researchers can share information on siRNA design and use.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Studies have shown that genetic and sex differences strongly influence gene expression in mice. Given the diversity and complexity of transcripts produced by alternative splicing, we sought to use ...microarrays to establish the extent of variation found in mouse strains and genders. Here, we surveyed the effect of strain and sex on liver gene and exon expression using male and female mice from three different inbred strains.
71 liver RNA samples from three mouse strains - DBA/2J, C57BL/6J and C3H/HeJ - were profiled using a custom-designed microarray monitoring exon and exon-junction expression of 1,020 genes representing 9,406 exons. Gene expression was calculated via two different methods, using the 3'-most exon probe ("3' gene expression profiling") and using all probes associated with the gene ("whole-transcript gene expression profiling"), while exon expression was determined using exon probes and flanking junction probes that spanned across the neighboring exons ("exon expression profiling"). Widespread strain and sex influences were detected using a two-way Analysis of Variance (ANOVA) regardless of the profiling method used. However, over 90% of the genes identified in 3' gene expression profiling or whole transcript profiling were identified in exon profiling, along with 75% and 38% more genes, respectively, showing evidence of differential isoform expression. Overall, 55% and 32% of genes, respectively, exhibited strain- and sex-bias differential gene or exon expression.
Exon expression profiling identifies significantly more variation than both 3' gene expression profiling and whole-transcript gene expression profiling. A large percentage of genes that are not differentially expressed at the gene level demonstrate exon expression variation suggesting an influence of strain and sex on alternative splicing and a need to profile expression changes at sub-gene resolution.
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Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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