Dually Active Polycation/miRNA Nanoparticles for the Treatment of Fibrosis in Alcohol-Associated Liver Disease.

Alcohol-associated liver disease (AALD) is a major cause of liver disorders worldwide. Current treatment options are limited, especially for AALD-associated fibrosis. Promising approaches include RNA interference for miR-155 overexpression in Kupffer cells (KCs), as well as the use of CXCR4 antagonists that inhibit the activation of hepatic stellate cells (HSCs) through the CXCL12/CXCR4 axis. The development of dual-functioning nanoparticles for the effective delivery of antifibrotic RNA together with a CXCR4 inhibitor thus promises to improve the treatment of AALD fibrosis. In this study, cholesterol-modified polymeric CXCR4 inhibitor (Chol-PCX) was synthesized and used to encapsulate anti-miR-155 or non-coding (NC) miRNA in the form of Chol-PCX/miRNA nanoparticles. The results indicate that the nanoparticles induce a significant miR-155 silencing effect both in vitro and in vivo. Treatment with the Chol-PCX/anti-miR-155 particles in a model of moderate alcohol consumption with secondary liver insult resulted in a significant reduction in aminotransferase enzymes as well as collagen content in the liver parenchyma. Overall, our data support the use of Chol-PCX as a carrier for anti-miR-155 for the combined therapeutic inhibition of CXCR4 and miR-155 expression as a way to improve fibrotic damage in the liver.

Collagen fiber regulation in human pediatric aortic valve development and disease.

Congenital aortic valve stenosis (CAVS) affects up to 10% of the world population without medical therapies to treat the disease. New molecular targets are continually being sought that can halt CAVS progression. Collagen deregulation is a hallmark of CAVS yet remains mostly undefined. Here, histological studies were paired with high resolution accurate mass (HRAM) collagen-targeting proteomics to investigate collagen fiber production with collagen regulation associated with human AV development and pediatric end-stage CAVS (pCAVS). Histological studies identified collagen fiber realignment and unique regions of high-density collagen in pCAVS. Proteomic analysis reported specific collagen peptides are modified by hydroxylated prolines (HYP), a post-translational modification critical to stabilizing the collagen triple helix. Quantitative data analysis reported significant regulation of collagen HYP sites across patient categories. Non-collagen type ECM proteins identified (26 of the 44 total proteins) have direct interactions in collagen synthesis, regulation, or modification. Network analysis identified BAMBI (BMP and Activin Membrane Bound Inhibitor) as a potential upstream regulator of the collagen interactome. This is the first study to detail the collagen types and HYP modifications associated with human AV development and pCAVS. We anticipate that this study will inform new therapeutic avenues that inhibit valvular degradation in pCAVS and engineered options for valve replacement.

Spinal Cord Stimulation for the Treatment of Chronic Pain Reduces Opioid Use and Results in Superior Clinical Outcomes When Used Without Opioids.

BACKGROUND: Chronic pain causes a significant burden to the US health care system, is difficult to treat, and remains a significant contributor to increased opioid use in the United States. Spinal cord stimulation (SCS) has been FDA approved for the treatment of chronic pain. OBJECTIVE: To evaluate the hypothesis that SCS reduces opioid use, and alone maintains clinical outcome measures of pain and psychosocial determinants of health. METHODS: In this prospective cohort study, we evaluated 86 patients undergoing SCS surgery for the treatment of chronic pain between September 2012 and August 2015. Preoperatively and postoperatively, patients completed the Numerical Rating Scale (NRS), McGill Pain Questionnaire (MPQ), Pain Catastrophizing Scale (PCS), Oswestry Disability Index (ODI), and Beck's Depression Inventory (BDI). VAS scores were retrospectively analyzed. RESULTS: Fifty-three patients used opioids before SCS implantation. The 33 nonusers had lower mean VAS, NRS, and ODI scores than both opioid groups at 1 yr and improved significantly at 1 yr on the VAS (P < .001), NRS (P < .001), MPQ (P = .002), PCS (P < .001), BDI (P = .04), and ODI (P = .002). After surgery, 41.5% remained opioids and 58.5% reduced/eliminated use. Discontinued (n = 29) or reduced (n = 2) use resulted in VAS, NRS, total MPQ, and ODI score reduction (P < .001, P = .002, P = .002, and P = .009 respectively). At 1 yr, survey scores in opioid users were unchanged. There was no difference between groups in revision or failure rates. CONCLUSION: Sixty-four percent of patients who were using opioids prior to SCS reduced (n = 2) or eliminated opioid use (n = 29) at 1 yr postoperatively. Patients who eliminated opioid use or never used opioids had superior clinical outcomes to those who continued use.

Distal Radius Fracture Outcomes and Rehabilitation.

Distal radius fractures account for nearly 1 of every 5 fractures in individuals aged 65 or older. Moreover, increased susceptibility to vertebral and hip fractures has been documented in patients a year after suffering a distal radius fracture. Although women are more susceptible to hip fractures, men experience a higher mortality rate in the 7 years following a distal radius fracture. Traditional approaches to distal radius fractures have included both surgical and nonsurgical treatments, with predominant complaints involving weakness, stiffness, and pain. Nonsurgical approaches include immobilization with or without reduction, whereas surgical treatments include dorsal spanning bridge plates, percutaneous pinning, external fixation, and volar plate fixation. The nature of the fracture will determine the best treatment option, and surgeons employ a multifactorial treatment approach that includes the patient's age, nature of injury, joint involvement, and displacement among other factors. Historically, closed reduction and percutaneous pinning have been the most popular approaches. However, volar plate fixation is quickly becoming a popular option as it minimizes tendon irritation, reduces immobilization time, and decreases risk of complication. The goal of treatment is to restore mobility, reduce pain, and improve functional outcomes following rehabilitation. The aim of this review is to summarize the most common treatments and importance of early referral to hand therapy to improve functional outcomes.

Amniotic fluid: the use of high-dimensional biology to understand fetal well-being.

Our aim was to review the use of high-dimensional biology techniques, specifically transcriptomics, proteomics, and metabolomics, in amniotic fluid to elucidate the mechanisms behind preterm birth or assessment of fetal development. We performed a comprehensive MEDLINE literature search on the use of transcriptomic, proteomic, and metabolomic technologies for amniotic fluid analysis. All abstracts were reviewed for pertinence to preterm birth or fetal maturation in human subjects. Nineteen articles qualified for inclusion. Most articles described the discovery of biomarker candidates, but few larger, multicenter replication or validation studies have been done. We conclude that the use of high-dimensional systems biology techniques to analyze amniotic fluid has significant potential to elucidate the mechanisms of preterm birth and fetal maturation. However, further multicenter collaborative efforts are needed to replicate and validate candidate biomarkers before they can become useful tools for clinical practice. Ideally, amniotic fluid biomarkers should be translated to a noninvasive test performed in maternal serum or urine.

Gentamicin rapidly inhibits mitochondrial metabolism in high-frequency cochlear outer hair cells.

Aminoglycosides (AG), including gentamicin (GM), are the most frequently used antibiotics in the world and are proposed to cause irreversible cochlear damage and hearing loss (HL) in 1/4 of the patients receiving these life-saving drugs. Akin to the results of AG ototoxicity studies, high-frequency, basal turn outer hair cells (OHCs) preferentially succumb to multiple HL pathologies while inner hair cells (IHCs) are much more resilient. To determine if endogenous differences in IHC and OHC mitochondrial metabolism dictate differential sensitivities to AG-induced HL, IHC- and OHC-specific changes in mitochondrial reduced nicotinamide adenine dinucleotide (NADH) fluorescence during acute (1 h) GM treatment were compared. GM-mediated decreases in NADH fluorescence and succinate dehydrogenase activity were observed shortly after GM application. High-frequency basal turn OHCs were found to be metabolically biased to rapidly respond to alterations in their microenvironment including GM and elevated glucose exposures. These metabolic biases may predispose high-frequency OHCs to preferentially produce cell-damaging reactive oxygen species during traumatic challenge. Noise-induced and age-related HL pathologies share key characteristics with AG ototoxicity, including preferential OHC loss and reactive oxygen species production. Data from this report highlight the need to address the role of mitochondrial metabolism in regulating AG ototoxicity and the need to illuminate how fundamental differences in IHC and OHC metabolism may dictate differences in HC fate during multiple HL pathologies.

MicroRNA-183 family expression in hair cell development and requirement of microRNAs for hair cell maintenance and survival.

MicroRNAs (miRNAs) post-transcriptionally repress complementary target gene expression and can contribute to cell differentiation. The coordinate expression of miRNA-183 family members (miR-183, miR-96, and miR-182) has been demonstrated in sensory cells of the mouse inner ear and other vertebrate sensory organs. To further examine hair cell miRNA expression in the mouse inner ear, we have analyzed miR-183 family expression in wild type animals and various mutants with defects in neurosensory development. miR-183 family member expression follows neurosensory cell specification, exhibits longitudinal (basal-apical) gradients in maturating cochlear hair cells, and is maintained in sensory neurons and most hair cells into adulthood. Depletion of hair cell miRNAs resulting from Dicer1 conditional knockout (CKO) in Atoh1-Cre transgenic mice leads to more disparate basal-apical gene expression profiles and eventual hair cell loss. Results suggest that hair cell miRNAs subdue cochlear gradient gene expression and are required for hair cell maintenance and survival.

A flow cytometric assay to quantify in vivo bacterial uptake by alveolar macrophages.

Our laboratory has developed a flow cytometric assay to quantify alveolar macrophage (Mcapital EF, Cyrillic) phagocytosis of bacteria within a live animal. Mcapital EF, Cyrillics collected by bronchoalveolar lavage from rats infected transtracheally with Syto 9-labeled bacteria are fluorescently labeled for identification and analyzed by flow cytometry to quantify their bacterial uptake.

Salmeterol/fluticasone propionate versus fluticasone propionate plus montelukast: a cost-effective comparison for asthma.

INTRODUCTION AND OBJECTIVE: Asthma, owing to its chronic nature, is associated with a substantial economic burden. Healthcare providers need to compare the cost effectiveness of alternative asthma treatment options to ensure that they obtain the best value for money from the resources they control. The objective of the current study was to compare the cost effectiveness of salmeterol/fluticasone propionate in combination with fluticasone propionate plus montelukast in patients with symptomatic asthma uncontrolled with inhaled corticosteroid (ICS) monotherapy. STUDY DESIGN AND METHODS: Direct healthcare resource data were prospectively collected during a double-blind, randomized, 12-week clinical study of inhaled salmeterol/fluticasone propionate 50/100 microg twice daily (n = 356) and inhaled fluticasone propionate 100 microg twice daily plus oral montelukast 10mg daily (n = 369). Resources were costed in Dutch guilders (NLG) from the perspective of The Netherlands healthcare system using 1999/2000 prices, but have been presented in US dollars and euros. The primary effectiveness measure was the proportion of successfully treated weeks (based on mean morning PEF values). Secondary measures were episode-free days, symptom-free days, and symptom-free nights. RESULTS: Salmeterol/fluticasone propionate was more effective than fluticasone propionate plus montelukast as measured by the proportion of successfully treated weeks mean 63.3% vs 39.0%; median difference 25%; p < 0.001). Salmeterol/fluticasone propionate was also more effective than fluticasone propionate plus montelukast according to the secondary effectiveness measures. The mean total direct daily healthcare costs per patient were 16% higher with fluticasone propionate plus montelukast than with salmeterol/fluticasone propionate mainly due to higher drug costs in the former group (2.25 US dollars vs 1.94; 1.92 euro vs 1.66, respectively; the NLG was fixed against the euro at a rate of 1 euro = NLG2.2 on 31 December 1998; 1 US dollars = NLG1.883, June 2003; 1 US dollars= 0.848 euro, June 2003). Incremental cost-effectiveness analyses showed that salmeterol/fluticasone propionate was dominant over fluticasone propionate plus montelukast and sensitivity analyses showed these results to be robust. CONCLUSION: Salmeterol/fluticasone propionate is a more cost-effective treatment option than fluticasone propionate plus montelukast for patients with symptomatic asthma uncontrolled by ICS.

Selective expression of beta tubulin isotypes in gerbil vestibular sensory epithelia and neurons.

The seven mammalian isotypes of beta tubulin are strikingly similar in amino acid sequence. The differences in isotypic sequence, although small, are nonetheless conserved in evolution, which suggests that they may confer distinct functional roles. If so, such roles should be reflected in the selective expression of isotypes by cell type, or even in the sorting of isotypes to within-cell pools. Hair cells of the vestibular sensory epithelia each possess a kinocilium, a microtubule-based organelle that could represent a distinct microtubule compartment, separate from the extensive microtubule network in the soma. The afferent neurons that innervate the vestibular sensory epithelia may also be functionally divided into dendritic, somatic, and axonal compartments, each with its own complement of microtubules. We have examined the distribution of beta tubulin isotypes in gerbil vestibular epithelia using isotype-specific antibodies to four isotypes and indirect immunofluorescence. We found that hair cells selectively express betaI and betaIV tubulin, while supporting cells express betaI, betaII, and betaIV tubulin. However, no sorting of isotypes between somatic and kinocilia compartments was found in hair cells. Vestibular ganglion cells display three isotypes in the soma, axon, and terminal dendrite compartments (betaI, betaII, and betaIII tubulin), but only betaIII tubulin was found in calyceal nerve endings. The implication of these findings is that beta tubulin isotypes are not sorted to within-cell compartments in hair cells but are sorted in some vestibular neurons.

Requirement for the betaI and betaIV tubulin isotypes in mammalian cilia.

Nielsen et al., [2001: Curr Biol 11:529-533], based on studies in Drosophila, have proposed that beta tubulin in axonemal microtubules must contain a specific acidic seven amino acid sequence in its carboxyl terminus. In mammals, the two betaIV isotypes (betaIVa and betaIVb) contain that sequence. In order to test the application of this hypothesis to mammals, we have examined the expression of beta tubulin isotypes in four different ciliated tissues (trachea, ependyma, uterine tube, and testis) using isotype-specific antibodies and indirect immunofluorescence. We find that betaIV tubulin is present in all ciliated cell types examined, but so is betaI tubulin. Taken together with recent studies that show that betaI and betaIV tubulin are both present in the cilia of vestibular hair cells, olfactory neurons, and nasal respiratory epithelial cells, we propose that both betaI tubulin and betaIV tubulin may be required for axonemal structures in mammals.

Cell type-specific reduction of beta tubulin isotypes synthesized in the developing gerbil organ of Corti.

There are seven isotypic forms of the microtubule protein beta tubulin in mammals, but not all isotypes are synthesized in every cell type. In the adult organ of Corti, each of the five major cell types synthesizes a different subset of isotypes. Inner hair cells synthesize only betaI and betaII tubulin, while outer hair cells make betaI and betaIV tubulin. Only betaII and betaIV tubulin are found in inner and outer pillar cells, while betaI, betaII, and betaIV tubulin are present in Deiters cells, and betaI, betaII and betaIII tubulin are found in organ of Corti dendrites. During post-natal organ of Corti development in the gerbil, microtubules are elaborated in an orderly temporal sequence beginning with hair cells, followed by pillar cells and Deiters cells. Using beta tubulin isotype-specific antibodies, we show that, in the gerbil cochlea, the same three isotypes are present in each cell type at birth, and that a cell type-specific reduction in the isotypes synthesized occurs in hair cells and pillar cells at an unusually late stage in development. No beta tubulin isotypes were detected in mature afferent dendrites, but we show that this is because few microtubules are present in mature dendrites. In addition, we show that primary cilia in inner hair cells, a feature of early development, persist much later than previously reported. The findings represent the first description of developmental cell type-specific reductions in tubulin isotypes in any system.

Differential synthesis of beta-tubulin isotypes in gerbil nasal epithelia.

Compartmentalization of beta-tubulin isotypes within cells according to function was examined in gerbil olfactory and respiratory epithelia by using specific antibodies to four beta-tubulin isotypes (beta(I), beta(II), beta(III), and beta(IV)). Isotype synthesis was cell-type-specific, but the localization of the isotypes was not compartmentalized. All four isotypes were found in the cilia, dendrites, somata, and axons of olfactory neurons. Only two isotypes (beta(I) and beta(IV)) were present in the cilia of nasal respiratory epithelial cells. The beta(IV) isotype, thought to be an essential component of cilia, was present in olfactory neurons and respiratory epithelial cells, which are ciliated, but was not found in basal cells (the stem cells of olfactory sensory neurons, which have no cilia). Olfactory neurons therefore do not synthesize beta(IV)-tubulin until they mature, when functioning cilia are also elaborated. The failure to observe compartmentalization of beta-tubulin isotypes in olfactory neurons sheds new light on potential functions of the beta-tubulin isotypes.