To Cross or Not to Cross: Using MRI-Histology to Characterize Dense Collagenous Plaque in Critical Limb Ischemia.

Peripheral artery disease (PAD) is caused by atherosclerotic buildup in the lower extremities, leading to obstruction and inadequate perfusion to the peripheral vasculature. Impenetrable plaques initially treated with percutaneous vascular intervention (PVI) have led to worse secondary bypass outcomes and amputation in patients. In this case report, we discuss the importance of using magnetic resonance imaging (MRI) histology in PVI planning in a patient with critical limb ischemia. PVI attempts to recanalize the limb failed because of an impenetrable occlusion in the popliteal artery that was not identified on routine preoperative imaging. Subsequent bypass occluded multiple times eventually requiring an above-knee amputation. An MRI-histology protocol-using ultrashort echo time (UTE) and T2-weighted (T2W) sequences-that was performed prior to the index PVI identified the occlusion as a dense collagen plaque. Histology analysis of the amputated specimen confirmed the MRI finding. This imaging modality offers a novel approach to characterize plaque composition and morphology, thereby identifying lesions at greatest risk of PVI failure and potentially playing an important role in selecting the right candidates for an endovascular-first approach.

Catheter-Directed Interventions for the Treatment of Lower Extremity Deep Vein Thrombosis.

Lower extremity deep vein thrombosis (DVT) leads to significant morbidity including pain, swelling, and difficulty walking in the affected limb. If left untreated, DVT increases the risk of pulmonary embolism (PE), recurrent venous thromboembolism (VTE), and post thrombotic syndrome (PTS). The objective of this review was to identify catheter-directed interventions and their success rates for the treatment of lower extremity DVT. A comprehensive search of current and emerging catheter-directed interventions for lower extremity DVT treatment was conducted in PubMed and Google Scholar. Clinical trials, retrospective and prospective observational studies, and case reports were identified to classify percutaneous mechanical thrombectomy (PMT), catheter-directed thrombolysis (CDT), and pharmacomechanical CDT (PCDT) devices based on their mechanism of action and indication of use. Catheter-directed interventions such as PMT, CDT, and PCDT offer an alternative therapeutic strategy for DVT management, particularly in patients with limb-threatening conditions and absolute contraindications to anticoagulants. Currently, there are limited guidelines for the use of mechanical and pharmacomechanical devices because of the lack of clinical evidence available for their use in treatment. Future studies are required to determine the short and long-term effects of using catheter-directed interventions as well as their effectiveness in treating acute versus subacute and chronic DVT.

Increasing statin prescription rates to prevent cardiovascular disease among high-risk populations: a quality improvement intervention centred on a novel interactive tool.

Statins are indicated for primary and secondary prevention of atherosclerotic cardiovascular disease (ASCVD). Our previous study of 1042 consecutive patient encounters at our large urban academic institution found that one in five patients were not prescribed an appropriate statin therapy. Only one-third of patients had follow-up cholesterol levels ordered to monitor treatment efficacy. In order to improve adherence to cholesterol guidelines at our institution, a quality improvement project was undertaken. We implemented interventions over a 4-month period to improve statin prescription rates: (a) development of an online interactive tool, (b) physician education on updated cholesterol guidelines and utilisation of the tool, (c) display of guideline summary in the workspace and (d) a documentation reminder in the electronic health record. We randomly selected encounter dates, from which 622 consecutive patient encounters were analysed. The primary outcome measures were prescription rates of statins, documentation of a 10-year ASCVD risk score and follow-up cholesterol levels ordered to monitor treatment efficacy. Out of the 622 patient encounters, 232 met statin indication. In this post-intervention group, statin prescription rates improved when compared with the pre-intervention group (90.5% vs 82.3%, p=0.006). Among patients who met statin indication solely via a 10-year ASCVD risk score ≥7.5%, there was an increase in documentation of the calculated 10-year ASCVD risk score (72.3% vs 57.8%; p=0.039) and in statin prescription rate (90.8% vs 67.6%; p<0.001). In addition, there was an increase in follow-up cholesterol levels ordered in all patients included in our study who met statin indication (64.1% vs 33.3%; p<0.001). Our quality improvement project showed higher rates of statin prescription, 10-year ASCVD risk score documentation and treatment monitoring after multiple interventions, centred on an easily accessible online interactive tool.

A Rare Presentation of Infective Endocarditis Due to Serratia marcescens.

Serratia marcescens is an opportunistic organism that can commonly cause respiratory tract infections in immunocompromised individuals. It has also been shown to cause urinary tract infections and soft tissue infections. It has several virulence factors including fimbriae-like adhesions that allow for surface attachment and biofilm formation to increase the likelihood of infections in humans. However, it has rarely been shown to cause infective endocarditis but has an increased mortality compared to the usual microbial agents associated with it (Staphylococcus and Streptococcus). Therefore, a high index of suspicion is necessary to accurately diagnose and treat patients at risk. Most published cases of S. marcescens endocarditis show that almost all described patients had chronic medical conditions or cardiovascular abnormalities. Furthermore, treatment has become difficult as S. marcescens has been shown to exhibit antibacterial resistance with beta-lactamase production. Here, we present a complicated case of S. marcescens pneumonia and infective endocarditis with a good prognosis. Our patient had a rapid onset of complications (i.e. including joint infections, splenic abscesses, myositis, and septic arthritis), despite the initial benign presentation concerning for pneumonia. However, the patient had a favorable outcome due to the prompt work-up and treatment that was initiated. Therefore, S. marcescens bacteremia in a patient with risk factors should prompt further investigation with a thorough evaluation of source followed by immediate management. This case highlights the fastidious nature of S. marcescens. Further investigation needs to be done to elucidate the pathogenesis of the organism that can serve as a target for future therapeutic intervention.

Calcium Channels in the Heart: Disease States and Drugs.

Calcium ions are the major signaling ions in the cells. They regulate muscle contraction, neurotransmitter secretion, cell growth and migration, and the activity of several proteins including enzymes and ion channels and transporters. They participate in various signal transduction pathways, thereby regulating major physiological functions. Calcium ion entry into the cells is regulated by specific calcium channels and transporters. There are mainly six types of calcium channels, of which only two are prominent in the heart. In cardiac tissues, the two types of calcium channels are the L type and the T type. L-type channels are found in all cardiac cells and T-type are expressed in Purkinje cells, pacemaker and atrial cells. Both these types of channels contribute to atrioventricular conduction as well as pacemaker activity. Given the crucial role of calcium channels in the cardiac conduction system, mutations and dysfunctions of these channels are known to cause several diseases and disorders. Drugs targeting calcium channels hence are used in a wide variety of cardiac disorders including but not limited to hypertension, angina, and arrhythmias. This review summarizes the type of cardiac calcium channels, their function, and disorders caused by their mutations and dysfunctions. Finally, this review also focuses on the types of calcium channel blockers and their use in a variety of cardiac disorders.

Clostridial Mycotic Aneurysm Leading to Emphysematous Aortitis.

Mycotic aneurysms account for less than 5% of all aneurysms of the aorta, with most cases linked to infection with either Staphylococcus or Salmonella species. Emphysematous aortitis is a rare consequence of mycotic aneurysms and is associated with high morbidity and mortality. It typically occurs from infection superimposed on already damaged endothelium, which is commonly seen in conditions such as atherosclerosis. This report discusses the presentation and relevant imaging findings of a unique case of emphysematous aortitis from Clostridial infection of the thoracic aorta. The patient was a 66-year-old male with a past medical history of end-stage renal disease, arteriovenous fistula for dialysis, hypertension, and diabetes, who presented with tachycardia and tachypnea. Computed tomography of the chest showed inflammatory changes of the thoracic aorta with gas bubbles along the aortic wall, and post-mortem aortic tissue cultures were positive for Clostridium innocuum. Although emphysematous aortitis is rare, the radiographic findings are strikingly characteristic and should prompt immediate and aggressive management.

Stenotrophomonas maltophilia: An Emerging Pathogen of the Respiratory Tract.

BACKGROUND Stenotrophomonas maltophilia has the propensity to cause a plethora of opportunistic infections in humans owing to biofilm formation and antibiotic resistance. It is often seen as a co-organism along with Pseudomonas aeruginosa. CASE REPORT A 70-year-old woman with several co-morbidities presented reporting hypoglycemia and dyspnea. An imaging study of the chest was suggestive of deterioration of pneumonia, with increased opacities. Initial respiratory cultures were negative, while subsequent repeat cultures revealed the growth of Stenotrophomonas maltophilia susceptible to trimethoprim plus sulfamethoxazole and levofloxacin. The patient had a poor prognosis and eventually died despite appropriate measures. CONCLUSIONS A decline in the clinical status of a patient such as ours makes it hard to quickly diagnose this organism correctly. Physicians should thus be cautious of Stenotrophomonas maltophilia-induced infection and more emphasis should be placed on appropriate treatment due to the emerging risk of antibiotic resistance.

BKCa (Slo) Channel Regulates Mitochondrial Function and Lifespan in Drosophila melanogaster.

BKCa channels, originally discovered in Drosophila melanogaster as slowpoke (slo), are recognized for their roles in cellular and organ physiology. Pharmacological approaches implicated BKCa channels in cellular and organ protection possibly for their ability to modulate mitochondrial function. However, the direct role of BKCa channels in regulating mitochondrial structure and function is not deciphered. Here, we demonstrate that BKCa channels are present in fly mitochondria, and slo mutants show structural and functional defects in mitochondria. slo mutants display an increase in reactive oxygen species and the modulation of ROS affected their survival. We also found that the absence of BKCa channels reduced the lifespan of Drosophila, and overexpression of human BKCa channels in flies extends life span in males. Our study establishes the presence of BKCa channels in mitochondria of Drosophila and ascertains its novel physiological role in regulating mitochondrial structural and functional integrity, and lifespan.

Low cerebral blood flow is a non-invasive biomarker of neuroinflammation after repetitive mild traumatic brain injury.

Previous work has shown that non-invasive optical measurement of low cerebral blood flow (CBF) is an acute biomarker of poor long-term cognitive outcome after repetitive mild traumatic brain injury (rmTBI). Herein, we explore the relationship between acute cerebral blood flow and underlying neuroinflammation. Specifically, because neuroinflammation is a driver of secondary injury after TBI, we hypothesized that both glial activation and inflammatory signaling are associated with acute CBF and, by extension, with long-term cognitive outcome after rmTBI. To test this hypothesis, cortical CBF was non-invasively measured in anesthetized mice 4 h after 3 repetitive closed head injuries spaced once-daily, at which time brains were collected. Right hemispheres were fixed for immunohistochemical staining for glial activation markers Iba1 and GFAP while left hemispheres were used to quantify Iba1 and GFAP expression via Western blot as well as 32 cytokines and 21 phospho-proteins in the MAPK, PI3K/Akt, and NF-κB pathways using a Luminex multiplexed immunoassay. N = 8/7 injured/sham C57/black-6 adult male mice were studied. Within the injured group, CBF inversely correlated with Iba1 expression (R = -0.86, p < .01). Further, partial least squares regression analysis revealed significant correlations between CBF and expression of multiple pro-inflammatory cytokines, including RANTES and IL-17. Finally, within the injured group, phosphorylation of specific signals in the MAPK and NF-κB intracellular signaling pathways (e.g., p38 MAPK and NF-κB) were significantly positively correlated with Iba1. In total, our data indicate that acute cerebral blood flow after rmTBI is a biomarker of underlying neuroinflammatory pathology.

Inhibition of BKCa negatively alters cardiovascular function.

Large conductance calcium and voltage-activated potassium channels (BKCa ) are transmembrane proteins, ubiquitously expressed in the majority of organs, and play an active role in regulating cellular physiology. In the heart, BKCa channels are known to play a role in regulating the heart rate and protect it from ischemia-reperfusion injury. In vascular smooth muscle cells, the opening of BKCa channels results in membrane hyperpolarization which eventually results in vasodilation mediated by a reduction in Ca2+ influx due to the closure of voltage-dependent Ca2+ channels. Ex vivo studies have shown that BKCa channels play an active role in the regulation of the function of the majority of blood vessels. However, in vivo role of BKCa channels in cardiovascular function is not completely deciphered. Here, we have evaluated the rapid in vivo role of BKCa channels in regulating the cardiovascular function by using two well-established, rapid-acting, potent blockers, paxilline and iberiotoxin. Our results show that BKCa channels are actively involved in regulating the heart rate, the function of the left and right heart as well as major vessels. We also found that the effect on BKCa channels by blockers is completely reversible, and hence, BKCa channels can be exploited as potential targets for clinical applications for modulating heart rate and cardiac contractility.

Heme and hemoglobin suppress amyloid ß-mediated inflammatory activation of mouse astrocytes.

Glial immune activity is a key feature of Alzheimer's disease (AD). Given that the blood factors heme and hemoglobin (Hb) are both elevated in AD tissues and have immunomodulatory roles, here we sought to interrogate their roles in modulating ß-amyloid (Aß)-mediated inflammatory activation of astrocytes. We discovered that heme and Hb suppress immune activity of primary mouse astrocytes by reducing expression of several proinflammatory cytokines (e.g. RANTES (regulated on activation normal T cell expressed and secreted)) and the scavenger receptor CD36 and reducing internalization of Aß(1-42) by astrocytes. Moreover, we found that certain soluble (>75-kDa) Aß(1-42) oligomers are primarily responsible for astrocyte activation and that heme or Hb association with these oligomers reverses inflammation. We further found that heme up-regulates phosphoprotein signaling in the phosphoinositide 3-kinase (PI3K)/Akt pathway, which regulates a number of immune functions, including cytokine expression and phagocytosis. The findings in this work suggest that dysregulation of Hb and heme levels in AD brains may contribute to impaired amyloid clearance and that targeting heme homeostasis may reduce amyloid pathogenesis. Altogether, we propose heme as a critical molecular link between amyloid pathology and AD risk factors, such as aging, brain injury, and stroke, which increase Hb and heme levels in the brain.

Drosophila Voltage-Gated Calcium Channel α1-Subunits Regulate Cardiac Function in the Aging Heart.

Ion channels maintain numerous physiological functions and regulate signaling pathways. They are the key targets for cellular reactive oxygen species (ROS), acting as signaling switches between ROS and ionic homeostasis. We have carried out a paraquat (PQ) screen in Drosophila to identify ion channels regulating the ROS handling and survival in Drosophila melanogaster. Our screen has revealed that α1-subunits (D-type, T-type, and cacophony) of voltage-gated calcium channels (VGCCs) handle PQ-mediated ROS stress differentially in a gender-based manner. Since ROS are also involved in determining the lifespan, we discovered that the absence of T-type and cacophony decreased the lifespan while the absence of D-type maintained a similar lifespan to that of the wild-type strain. VGCCs are also responsible for electrical signaling in cardiac cells. The cardiac function of each mutant was evaluated through optical coherence tomography (OCT), which revealed that α1-subunits of VGCCs are essential in maintaining cardiac rhythmicity and cardiac function in an age-dependent manner. Our results establish specific roles of α1-subunits of VGCCs in the functioning of the aging heart.

Expression and Activation of BKCa Channels in Mice Protects Against Ischemia-Reperfusion Injury of Isolated Hearts by Modulating Mitochondrial Function.

Aims: Activation and expression of large conductance calcium and voltage-activated potassium channel (BKCa) by pharmacological agents have been implicated in cardioprotection from ischemia-reperfusion (IR) injury possibly by regulating mitochondrial function. Given the non-specific effects of pharmacological agents, it is not clear whether activation of BKCa is critical to cardioprotection. In this study, we aimed to decipher the mechanistic role of BKCa in cardioprotection from IR injury by genetically activating BKCa channels. Methods and Results: Hearts from adult (3 months old) wild-type mice (C57/BL6) and mice expressing genetically activated BKCa (Tg-BKCa R207Q, referred as Tg-BKCa) along with wild-type BKCa were subjected to 20 min of ischemia and 30 min of reperfusion with or without ischemic preconditioning (IPC, 2 times for 2.5 min interval each). Left ventricular developed pressure (LVDP) was recorded using Millar's Mikrotip® catheter connected to ADInstrument data acquisition system. Myocardial infarction was quantified by 2,3,5-triphenyl tetrazolium chloride (TTC) staining. Our results demonstrated that Tg-BKCa mice are protected from IR injury, and BKCa also contributes to IPC-mediated cardioprotection. Cardiac function parameters were also measured by echocardiography and no differences were observed in left ventricular ejection fraction, fractional shortening and aortic velocities. Amplex Red® was used to assess reactive oxygen species (ROS) production in isolated mitochondria by spectrofluorometry. We found that genetic activation of BKCa reduces ROS after IR stress. Adult cardiomyocytes and mitochondria from Tg-BKCa mice were isolated and labeled with Anti-BKCa antibodies. Images acquired via confocal microscopy revealed localization of cardiac BKCa in the mitochondria. Conclusions: Activation of BKCa is essential for recovery of cardiac function after IR injury and is likely a factor in IPC mediated cardioprotection. Genetic activation of BKCa reduces ROS produced by complex I and complex II/III in Tg-BKCa mice after IR, and IPC further decreases it. These results implicate BKCa-mediated cardioprotection, in part, by reducing mitochondrial ROS production. Localization of Tg-BKCa in adult cardiomyocytes of transgenic mice was similar to BKCa in wild-type mice.

Optimization of Non-Thermal Plasma Treatment in an In Vivo Model Organism.

Non-thermal plasma is increasingly being recognized for a wide range of medical and biological applications. However, the effect of non-thermal plasma on physiological functions is not well characterized in in vivo model systems. Here we use a genetically amenable, widely used model system, Drosophila melanogaster, to develop an in vivo system, and investigate the role of non-thermal plasma in blood cell differentiation. Although the blood system in Drosophila is primitive, it is an efficient system with three types of hemocytes, functioning during different developmental stages and environmental stimuli. Blood cell differentiation in Drosophila plays an essential role in tissue modeling during embryogenesis, morphogenesis and also in innate immunity. In this study, we optimized distance and frequency for a direct non-thermal plasma application, and standardized doses to treat larvae and adult flies so that there is no effect on the viability, fertility or locomotion of the organism. We discovered that at optimal distance, time and frequency, application of plasma induced blood cell differentiation in the Drosophila larval lymph gland. We articulate that the augmented differentiation could be due to an increase in the levels of reactive oxygen species (ROS) upon non-thermal plasma application. Our studies open avenues to use Drosophila as a model system in plasma medicine to study various genetic disorders and biological processes where non-thermal plasma has a possible therapeutic application.

Data supporting characterization of CLIC1, CLIC4, CLIC5 and DmCLIC antibodies and localization of CLICs in endoplasmic reticulum of cardiomyocytes.

Chloride intracellular channel (CLICs) proteins show 60-70% sequence identity to each other, and exclusively localize to the intracellular organelle membranes and cytosol. In support of our recent publication, "Molecular identity of cardiac mitochondrial chloride intracellular channel proteins" (Ponnalagu et al., 2016) [1], it was important to characterize the specificity of different CLIC paralogs/ortholog (CLIC1, CLIC4, CLIC5 and DmCLIC) antibodies used to decipher their localization in cardiac cells. In addition, localization of CLICs in the other organelles such as endoplasmic reticulum (ER) of cardiomyocytes was established. This article also provides data on the different primers used to show the relative abundance of CLIC paralogs in cardiac tissue and the specificity of the various CLIC antibodies used. We demonstrate that the predominant CLICs in the heart, namely CLIC1, CLIC4 and CLIC5, show differential distribution in endoplasmic reticulum. CLIC1 and CLIC4 both show co-localization to the endoplasmic reticulum whereas CLIC5 does not.

Molecular identity of cardiac mitochondrial chloride intracellular channel proteins.

Emerging evidences demonstrate significance of chloride channels in cardiac function and cardioprotection from ischemia-reperfusion (IR) injury. Unlike mitochondrial potassium channels sensitive to calcium (BKCa) and ATP (KATP), molecular identity of majority of cardiac mitochondrial chloride channels located at the inner membrane is not known. In this study, we report the presence of unique dimorphic chloride intracellular channel (CLIC) proteins namely CLIC1, CLIC4 and CLIC5 as abundant CLICs in the rodent heart. Further, CLIC4, CLIC5, and an ortholog present in Drosophila (DmCLIC) localize to adult cardiac mitochondria. We found that CLIC4 is enriched in the outer mitochondrial membrane, whereas CLIC5 is present in the inner mitochondrial membrane. Also, CLIC5 plays a direct role in regulating mitochondrial reactive oxygen species (ROS) generation. Our study highlights that CLIC5 is localized to the cardiac mitochondria and directly modulates mitochondrial function.