Citations

Background— Cardiac dysfunction in failing hearts of human patients and animal models is associated with both microtubule densification and transverse-tubule (T-tubule) remodeling. Our objective was to investigate whether microtubule densification contributes to T-tubule remodeling and excitation–contraction coupling dysfunction in heart disease. Methods and Results— In a mouse model of pressure overload–induced cardiomyopathy by transaortic banding, colchicine, a microtubule depolymerizer, significantly ameliorated T-tubule remodeling and cardiac dysfunction. In cultured cardiomyocytes, microtubule depolymerization with nocodazole or colchicine profoundly attenuated T-tubule impairment, whereas microtubule polymerization/stabilization with taxol accelerated T-tubule remodeling. In situ immunofluorescence of heart tissue sections demonstrated significant disorganization of junctophilin-2 (JP2), a protein that bridges the T-tubule and sarcoplasmic reticulum membranes, in transaortic banded hearts as well as in human failing hearts, whereas colchicine injection significantly preserved the distribution of JP2 in transaortic banded hearts. In isolated mouse cardiomyocytes, prolonged culture or treatment with taxol resulted in pronounced redistribution of JP2 from T-tubules to the peripheral plasma membrane, without changing total JP2 expression. Nocodazole treatment antagonized JP2 redistribution. Moreover, overexpression of a dominant-negative mutant of kinesin 1, a microtubule motor protein responsible for anterograde trafficking of proteins, protected against JP2 redistribution and T-tubule remodeling in culture. Finally, nocodazole treatment improved Ca 2+ handling in cultured myocytes by increasing the amplitude of Ca 2+ transients and reducing the frequency of Ca 2+ sparks. Conclusion— Our data identify a mechanistic link between microtubule densification and T-tubule remodeling and reveal microtubule-mediated JP2 redistribution as a novel mechanism for T-tubule disruption, loss of excitation–contraction coupling, and heart failure.

Abstract BACKGROUND Accurate evaluation of unclassified sequence variants in cancer predisposition genes is essential for clinical management and depends on a multifactorial analysis of clinical, genetic, pathologic, and bioinformatic variables and assays of transcript length and abundance. The integrity of assay data in turn relies on appropriate assay design, interpretation, and reporting. METHODS We conducted a multicenter investigation to compare mRNA splicing assay protocols used by members of the ENIGMA (Evidence-Based Network for the Interpretation of Germline Mutant Alleles) consortium. We compared similarities and differences in results derived from analysis of a panel of breast cancer 1, early onset (BRCA1) and breast cancer 2, early onset (BRCA2) gene variants known to alter splicing (BRCA1: c.135-1G>T, c.591C>T, c.594-2A>C, c.671-2A>G, and c.5467+5G>C and BRCA2: c.426-12_8delGTTTT, c.7988A>T, c.8632+1G>A, and c.9501+3A>T). Differences in protocols were then assessed to determine which elements were critical in reliable assay design. RESULTS PCR primer design strategies, PCR conditions, and product detection methods, combined with a prior knowledge of expected alternative transcripts, were the key factors for accurate splicing assay results. For example, because of the position of primers and PCR extension times, several isoforms associated with BRCA1, c.594-2A>C and c.671-2A>G, were not detected by many sites. Variation was most evident for the detection of low-abundance transcripts (e.g., BRCA2 c.8632+1G>A Δ19,20 and BRCA1 c.135-1G>T Δ5q and Δ3). Detection of low-abundance transcripts was sometimes addressed by using more analytically sensitive detection methods (e.g., BRCA2 c.426-12_8delGTTTT ins18bp). CONCLUSIONS We provide recommendations for best practice and raise key issues to consider when designing mRNA assays for evaluation of unclassified sequence variants.

In keeping with the stated goal of providing the surgical pathologist with tools to recognize abnormalities of the gastrointestinal (GI) tract due to drugs (AGIDS), in part II of this review we embark in a more organ-based description of AGIDS. Adequate space is given to the numerous adverse gastrointestinal effects of nonsteroidal anti-inflammatory drugs. Pill esophagitis, esophagitis dissecans, proton pump inhibitors’ effects, diaphragm disease, and the recently described effects of drugs such as olmesartan, mycophenolate, and of compounds such as yttrium-90 are highlighted among several others. The inclusion of drug effects in the differential diagnosis of “conventional” diseases (such as gastric antral vascular ectasia, graft-versus-host disease, ischemic colitis, acute colitis, collagenous enteritis, inflammatory bowel disease) is underscored to avoid sometimes significant diagnostic pitfalls. We reiterate the message of the necessary collaboration between pathologist and clinician in the recognition of these entities to provide the best patient care.

Abnormalities of the gastrointestinal (GI) tract due to drugs (AGIDs) are numerous and have significant impact. The aim of this narrative review is to help the practicing surgical pathologist recognize selected AGIDs. The adverse drug effects presented were chosen with an emphasis on recent and significant pathological and clinical contributions. The selection was based on a thorough review of the PUBMED-based literature and on the authors’ opinions and experience. In the first part of the review, diagnostic abnormalities due to crystals (eg, iron, biphosphonates, nonsystemic drugs), mitosis arresting drugs (colchicine, taxanes), and biological agents, especially ipilimumab, are discussed. Some AGIDs’ histopathologic features can be easily recognized. It is however the clinical correlation that in many cases of AGIDs will provide the necessary support for a drug effect diagnosis. The identification of AGIDs requires heightened awareness of the medical team in which close collaboration of pathologists and clinicians cannot be overemphasized.

Abstract Hypoxia-inducible factor 1 (HIF-1) is a master transcription factor that controls cellular homeostasis. Although its activation benefits normal tissue, HIF-1 activation in tumors is a major risk factor for angiogenesis, therapeutic resistance, and poor prognosis. HIF-1 activity is usually suppressed under normoxic conditions because of rapid oxygen-dependent degradation of HIF-1α. Here, we show that, under normoxic conditions, HIF-1α is upregulated in tumor cells in response to doxorubicin, a chemotherapeutic agent used to treat many cancers. In addition, doxorubicin enhanced VEGF secretion by normoxic tumor cells and stimulated tumor angiogenesis. Doxorubicin-induced accumulation of HIF-1α in normoxic cells was caused by increased expression and activation of STAT1, the activation of which stimulated expression of iNOS and its synthesis of nitric oxide (NO) in tumor cells. Mechanistic investigations established that blocking NO synthesis or STAT1 activation was sufficient to attenuate the HIF-1α accumulation induced by doxorubicin in normoxic cancer cells. To our knowledge, this is the first report that a chemotherapeutic drug can induce HIF-1α accumulation in normoxic cells, an efficacy-limiting activity. Our results argue that HIF-1α-targeting strategies may enhance doxorubicin efficacy. More generally, they suggest a broader perspective on the design of combination chemotherapy approaches with immediate clinical impact. Cancer Res; 73(20); 6230–42. ©2013 AACR.

SummaryBackgroundNeuronal loss via apoptosis in CNS is the fundamental mechanism underlying various neurodegenerative diseases. Compounds with antiapoptotic property might have therapeutic effects for these diseases. In this study, bis(propyl)‐cognitin (B3C), a novel dimer that possesses anti‐AChE and anti‐N‐methyl‐d‐aspartate receptor activities, was investigated for its neuroprotective effect on K+ deprivation‐induced apoptosis in cerebellar granule neurons (CGNs).MethodsCerebellar granule neurons were switched to K+ deprived medium with or without B3C. 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium assay, fluorescein diacetate (FDA)/propidium iodide (PI) staining, Hoechst staining, and DNA laddering assays were applied to detect cytotoxicity and apoptosis. Additionally, the expression of p‐VEGFR‐2, p‐Akt, p‐glycogen synthase kinase 3β (GSK3β), and p‐extracellular signal‐regulated kinase (ERK) was examined in CGNs.ResultsSwitching CGNs to K+ deprived medium resulted in remarkable apoptosis, which could be substantially blocked by B3C treatment (IC50, 0.37 μM). Moreover, a rapid decrease in p‐Tyr1054‐VEGFR‐2 was observed after the switch. B3C significantly reversed the inhibition of p‐Tyr1054‐VEGFR‐2 as well as Akt and ERK pathways. VEGFR‐2 inhibitor PTK787/ZK222584, as well as PI3‐K inhibitor LY294002 and MEK inhibitor PD98059, each abolished the neuroprotective effect of B3C.ConclusionsOur results demonstrate that B3C blocks K+ deprivation‐induced apoptosis in CGNs through regulating VEGFR‐2/Akt/GSK3β and VEGFR‐2/ERK signaling pathways, providing a molecular insight into the therapeutic potential of B3C for the treatment of neurodegenerative diseases.

Background: Decreased hip range of motion (ROM) is a common finding in patients with femoroacetabular impingement (FAI). Purpose: To report the prevalence of decreased hip ROM in asymptomatic adolescent athletes and to correlate examination findings to signs of FAI on radiographs and magnetic resonance imaging (MRI). Study Design: Cross-sectional study (prevalence); Level of evidence, 3. Methods: A total of 226 adolescent athletes presenting for state-mandated preparticipation physical examinations were assessed. Hip internal rotation was measured with the participant supine and the hip flexed to 90°. All participants with ≤10° of internal rotation were invited to return for standard radiographs and MRI of both hips. An age-matched control group, with >10° of internal rotation, underwent MRI examination only. Twenty-six athletes (13 study and 13 control) returned for clinical and radiographic examinations. Results: Nineteen athletes (34 hips, 8%) were found to have <10° of internal rotation. Eight athletes (13 hips, 3%) also had a positive anterior impingement sign. Thirteen of 19 athletes participated in the radiographic portion of the study. Of these 13 participants, 4 had limited internal rotation unilaterally, leaving 22 hips in the study group. Eight of 13 participants (15 hips, 68%) had a cam-type deformity evident on plain radiographs, and 4 participants (7 hips, 32%) had a positive radiographic crossover sign. The average α angle measured from radial MRI sequences was 58.1° in the study group versus 44.3° in the control group ( P < .001). Fifteen hips (68%) in the study group had abnormal MRI findings within the acetabular labrum or cartilage compared with 10 of 26 hips (38%) in the control group (odds ratio, 3.4; P = .078). Conclusion: Eight percent of asymptomatic teenagers had limited internal rotation on examination; 68% of these had radiographic findings suggestive of FAI. More than two thirds of these participants had evidence of asymptomatic hip pathological lesions on MRI.

Abstract The eosinophil-associated ribonuclease (Ear) family in the mouse consists of thirteen genes, eleven of which encode RNases that have physical/functional properties similar to the human Ears, eosinophil-derived neurotoxin and eosinophil cationic protein. The expression of Ear genes in the mouse is confined to sites of known eosinophilopoiesis, with the exception of the lung. Two Ear genes, Ear1 and Ear2, are predominantly expressed in the lungs of naive mice. Total Ear gene expression and RNase activity in bronchoalveolar lavage fluid increases significantly upon the induction of pulmonary inflammation using an ovalbumin (OVA) model of allergic sensitization and challenge. Interestingly, pulmonary Ear11 transcripts, which are absent in naive mice, accumulate as a consequence of OVA-mediated TH2 inflammation in the lung. The induction of Ear11 expression is dependent on the presence of T cells, in particular, CD4+ T lymphocytes. This effect is likely the result of the elaboration of TH2 cytokine levels, because pulmonary instillation of interleukin-4 or interleukin-13 induces the accumulation of Ear11 transcripts in naive animals. This study demonstrates that despite an allergen-mediated pulmonary eosinophilia and earlier studies showing that Ears are constituents of eosinophil secondary granules, alveolar macrophages are a significant source of these RNases in lungs of OVA-treated mice.