Calcitonin gene-related peptide and intermedin induce phosphorylation of p44/42 MAPK in primary human lymphatic endothelial cells in vitro.

Calcitonin gene-related peptide (CGRP) and adrenomedullin 2/intermedin (AM2/IMD) play important roles in several pathologies, including cardiovascular disease, migraine and cancer. The efficacy of drugs targeting CGRP signalling axis for the treatment of migraine patients is sometimes offset by side effects (e.g. inflammation and microvascular complications, including aberrant neovascularisation in the skin). Recent studies using animal models implicate CGRP in lymphangiogenesis and lymphatic vessel function. However, whether CGRP or AM2/IMD can act directly on lymphatic endothelial cells is unknown. Here, we found that CGRP and AM2/IMD induced p44/42 MAPK phosphorylation in a time- and dose-dependent manner in primary human dermal lymphatic endothelial cells (HDLEC) in vitro, and thus directly affected these cells. These new findings reveal CGRP and AM2/IMD as novel regulators of LEC biology and warrant further investigation of their roles in the context of pathologies associated with lymphatic function in the skin and other organs, and therapies targeting CGRP signalling axis.

Co-occurrence of Attention-Deficit/Hyperactivity Disorder and Septo-Optic Dysplasia: A Neurodevelopmental Case Report.

Septo-optic dysplasia (SOD) is a rare congenital disorder characterized by optic nerve hypoplasia, brain midline structure anomalies, and hypothalamic-pituitary axis hypoplasia. This case report aims to highlight the association between SOD and neurodevelopmental disorders, focusing on attention-deficit/hyperactivity disorder (ADHD) in addition to the well-established link with autism spectrum disorder (ASD). A six-year-old male diagnosed with SOD presented with behavioral concerns, including attention and impulse control issues. A comprehensive psychological evaluation confirmed the diagnosis of ADHD and ruled out ASD. Ophthalmological assessments were integral to understanding the patient's condition. This case underscores the importance of recognizing neurodevelopmental disorders in individuals with SOD, with a particular focus on the less common association with ADHD. The co-occurrence of these conditions underscores the complexity of neurodevelopmental disorders and the need for comprehensive evaluation and management. Collaboration between ophthalmologists and mental health specialists is crucial for addressing the diverse needs of these patients. Early identification and intervention for ADHD are essential for optimal developmental outcomes. This case underscores the necessity for further research to elucidate the relationship between SOD and ADHD, emphasizing the importance of holistic patient care and interdisciplinary collaboration in managing individuals with SOD spectrum conditions.

Amitriptyline-Induced Hyperprolactinemia in a Pediatric Patient.

Hyperprolactinemia is an endocrinological disorder that might arise from various physiologic or pathologic conditions, as well as from pharmacologic sources. These pharmacologic sources include antidepressants, antipsychotics, and dopamine receptor-blocking agents. Amitriptyline is classified as a tricyclic antidepressant. While it is FDA-approved primarily for the treatment of depression, amitriptyline also demonstrates efficacy in managing various other conditions, such as anxiety, post-traumatic stress disorder, insomnia, chronic and neuropathic pain, and migraine prevention. We present a case of a 10-year-old patient with a history of autism spectrum disorder (ASD), attention-deficit/hyperactivity disorder (ADHD), and migraine headaches who was incidentally found to have elevated prolactin levels while taking amitriptyline for migraine prophylaxis. While risperidone, an antipsychotic that can be used for ASD management, is commonly known to induce hyperprolactinemia, the association between amitriptyline and elevated prolactin is less frequently described in the literature. This case underscores the necessity for healthcare providers across various specialties to be aware of amitriptyline-induced hyperprolactinemia.

Quantitative proteomics reveals CLR interactome in primary human cells.

The G protein-coupled receptor (GPCR) calcitonin receptor-like receptor (CLR) mediates essential functions in several cell types and is implicated in cardiovascular pathologies, skin diseases, migraine, and cancer. To date, the network of proteins interacting with CLR ("CLR interactome") in primary cells, where this GPCR is expressed at endogenous (physiologically relevant) levels, remains unknown. To address this knowledge gap, we established a novel integrative methodological workflow/approach for conducting a comprehensive/proteome-wide analysis of Homo sapiens CLR interactome. We used primary human dermal lymphatic endothelial cells and combined immunoprecipitation utilizing anti-human CLR antibody with label-free quantitative nano LC-MS/MS and quantitative in situ proximity ligation assay. By using this workflow, we identified 37 proteins interacting with endogenously expressed CLR amongst 4902 detected members of the cellular proteome (by quantitative nano LC-MS/MS) and revealed direct interactions of two kinases and two transporters with this GPCR (by in situ proximity ligation assay). All identified interactors have not been previously reported as members of CLR interactome. Our approach and findings uncover the hitherto unrecognized compositional complexity of the interactome of endogenously expressed CLR and contribute to fundamental understanding of the biology of this GPCR. Collectively, our study provides a first-of-its-kind integrative methodological approach and datasets as valuable resources and robust platform/springboard for advancing the discovery and comprehensive characterization of physiologically relevant CLR interactome at a proteome-wide level in a range of cell types and diseases in future studies.

Ruxolitinib ameliorates progressive anemia and improves survival during episodes of emergency granulopoiesis in Fanconi C-/- mice.

Fanconi anemia (FA) is an inherited disorder of DNA repair with hematologic manifestations that range from anemia to bone marrow failure to acute myeloid leukemia. In a murine model of FA (Fancc-/- mice), we found bone marrow failure was accelerated by repeated attempts to induce emergency (stress) granulopoiesis, the process for granulocyte production during the innate immune response. Fancc-/- mice exhibited an impaired granulocytosis response and died with profound anemia during repeated challenge. In the current study, we found erythropoiesis and serum erythropoietin decreased in Fancc-/- and wild-type (Wt) mice as emergency granulopoiesis peaked. Serum erythropoietin returned to baseline during steady-state resumption, and compensatory proliferation of erythroid progenitors was associated with DNA damage and apoptosis in Fancc-/- mice, but not Wt mice. The erythropoietin receptor activates Janus kinase 2 (Jak2), and we found treatment of Fancc-/- mice with ruxolitinib (Jak1/2-inhibitor) decreased anemia, enhanced granulocytosis, delayed clonal progression and prolonged survival during repeated emergency granulopoiesis episodes. This was associated with a decrease in DNA damage and apoptosis in Fancc-/- erythroid progenitors during this process. Transcriptome analysis of these cells identified enhanced activity of pathways for metabolism of reactive oxygen species, and decreased apoptosis- and autophagy-related pathways, as major ruxolitinib-effects in Fancc-/- mice. In contrast, ruxolitinib influenced primarily pathways involved in proliferation and differentiation in Wt mice. Ruxolitinib is approved for treatment of myeloproliferative disorders and graft-versus-host disease, suggesting the possibility of translational use as a bone marrow protectant in FA.

Plasma TNFα and Unknown Factor/s Potentially Impede Erythroblast Enucleation Obstructing Terminal Maturation of Red Blood Cells in Burn Patients.

INTRODUCTION: In this study, using burn patient's peripheral blood mononuclear cells (PBMCs), we have shown that the Epo independent stage of terminal enucleation to reticulocyte formation is impeded in the presence of autologous plasma (BP). Furthermore, substitution with allogeneic control plasma (CP) from the healthy individual in place of BP rectified this enucleation defect. The exclusive role of burn microenvironment in late-stage erythropoiesis defect was further demarcated through control healthy human bone marrow cells cultured in the presence of CP, BP, and cytokines. METHODS: PBMCs and human bone marrow (huBM) were differentiated ex vivo to enucleated reticulocytes in the presence of required growth factors and 5% CP or BP. Effect of systemic mediators in burn microenvironment like IL-6, IL-15, and TNFα was also explored. Neutralization experiments were carried out by adding varying concentrations (25 ng-400 ng/mL) of Anti-TNFα Ab to either CP+TNFα or BP. RESULTS: Reticulocyte proportion and maturation index were significantly improved upon substituting BP with CP during differentiation of burn PBMCs. In the huBM ex vivo culture, addition of IL-6 and IL-15 to CP inhibited the proliferation stages of erythropoiesis, whereas TNFα supplementation caused maximum diminution at erythroblast enucleation stage. Supplementation with anti-TNFα in the BP showed significant but partial restoration in the enucleation process, revealing the possibility of other crucial microenvironmental factors that could impact RBC production in burn patients. CONCLUSION: Exogenous TNFα impairs late-stage erythropoiesis by blocking enucleation, but neutralization of TNFα in BP only partially restored terminal enucleation indicating additional plasma factor(s) impair(s) late-stage RBC maturation in burn patients.

Evolution of Distinct Responses to Low NAD+ Stress by Rewiring the Sir2 Deacetylase Network in Yeasts.

Evolutionary adaptation increases the fitness of a species in its environment. It can occur through rewiring of gene regulatory networks, such that an organism responds appropriately to environmental changes. We investigated whether sirtuin deacetylases, which repress transcription and require NAD+ for activity, serve as transcriptional rewiring points that facilitate the evolution of potentially adaptive traits. If so, bringing genes under the control of sirtuins could enable organisms to mount appropriate responses to stresses that decrease NAD+ levels. To explore how the genomic targets of sirtuins shift over evolutionary time, we compared two yeast species, Saccharomyces cerevisiae and Kluyveromyces lactis, that display differences in cellular metabolism and life cycle timing in response to nutrient availability. We identified sirtuin-regulated genes through a combination of chromatin immunoprecipitation and RNA expression. In both species, regulated genes were associated with NAD+ homeostasis, mating, and sporulation, but the specific genes differed. In addition, regulated genes in K. lactis were associated with other processes, including utilization of nonglucose carbon sources, detoxification of arsenic, and production of the siderophore pulcherrimin. Consistent with the species-restricted regulation of these genes, sirtuin deletion affected relevant phenotypes in K. lactis but not S. cerevisiae Finally, sirtuin-regulated gene sets were depleted for broadly conserved genes, consistent with sirtuins regulating processes restricted to a few species. Taken together, these results are consistent with the notion that sirtuins serve as rewiring points that allow species to evolve distinct responses to low NAD+ stress.

A Shift in Myeloid Cell Phenotype via Down Regulation of Siglec-1 in Island Macrophages of Bone Marrow Is Associated With Decreased Late Erythroblasts Seen in Anemia of Critical Illness.

Burn injury has been shown to significantly dampen erythropoiesis in both burn patients and in murine models. Our previous findings elucidated the erythropoietin independent defects in red cell development stages involving erythroid progenitor production and late stage erythroblast enucleation processes. We hypothesized that macrophages (MØ) in erythroblast islands (EBI) could be yet another roadblock impeding erythropoiesis following burn injury. Here we highlight that the methodology to study EBI can be achieved with single cell suspensions using a simple technique such as flow cytometry, as obtaining the central erythroblast island macrophages (EBIMØs) of interest is a delicate process. We elucidated the requisite of EBIMØ from the erythroblast as well as the MØ perspective. In addition to the primary erythropoiesis organ, the bone marrow (BM), spleens were also examined for extra-medullary erythropoiesis. Femurs and spleens were harvested from adult mice (B6D2F1) subjected to 15% total body surface area (TBSA) scald burn (B) or sham burn (S). Total bone marrow cells (TBM) and splenocytes were probed for total erythrons, early and late erythroblasts and EBIMØ by flow cytometry. There was only a marginal increase in the number of EBIMØ after burn, but, between the signatures of EBIMØ, Siglec-1 expression (MFI) was reduced by 40% in B with and a parallel 44% decrease in TBM erythrons in the BM. There were more (2.5-fold) EEBs and less LEBs (2.4-fold) per million TBM cells in B; with a corresponding decrease in Siglec-1 and Ly6G expressions in EBIMØ associated with EEB. Conversely, extra-medullary erythropoiesis was robust in spleens from B. Not only were the numbers of EBIMØs increased in B (p < 0.002), both EEBs and LEBs associated with EBIMØ were higher by 30 and 75%, respectively. Importantly, an increase in Siglec-1 and Vcam1 expressing F480+ splenic macrophages was observed after burn injury. Therefore, stagnant EEBs in the BM after burn injury could be due to low Siglec1 expressing EBIMØ, which perhaps impede their maturation into LEBs and reticulocytes. Repercussion of myeloid cell phenotype specific to BM after burn injury could plausibly account for a defective late stage RBC maturation resulting in anemia of critical illness. Summary Sentence: Characterization of erythroblast island macrophages (EBIMØ) in the bone marrow and spleen at different stages of erythropoiesis after burn injury.

Pre-Exposure to Chronic Unpredictable Stress Suppresses the Chemopreventive Potential of Aloe Vera (Av) Leaf Gel Against 7,12-Dimethylbenz(a)anthracene (DMBA) Induced Carcinogenesis.

The present study evaluates the topical application of aloe vera (Av) leaf gel as a protective natural product against 7,12-dimethylbenz(a)anthracene (DMBA)-induced skin lesions in Swiss albino mice and as an antioxidant for the systemic toxicity of DMBA in the presence and absence of chronic unpredictable stress (CUS). Animals were randomized into seven groups and sacrificed after 16 weeks of treatment. Av gel application along with DMBA + 12-O-tetradecanoylphorbol-13-acetate (TPA) was found to be effective in reducing tumor incidence, cumulative number of papillomas, tumor burden and tumor yield when compared to untreated groups. Furthermore, topical treatment with Av gel significantly increased the overall in vivo antioxidant status of mice. Conversely, lipid peroxidation levels were significantly decreased in skin and circulation. However, pre-exposure to CUS followed by DMBA + TPA + Av gel application reduced the chemopreventive efficacy of Av gel as evidenced by increased tumor incidence, tumor burden, tumor yield and MDA levels accompanied by decrease in the enzymatic and nonenzymatic antioxidants. These observations were further supported by the results of fluorescent studies and comet assay. The study demonstrates a reduction in the antioxidant and antitumor potential of Av gel in presence of CUS thereby, signifying the need of stress reduction during cancer chemopreventive trials.

Dynamins 2 and 3 control the migration of human megakaryocytes by regulating CXCR4 surface expression and ITGB1 activity.

Megakaryocyte (MK) migration from the bone marrow periosteal niche toward the vascular niche is a prerequisite for proplatelet extension and release into the circulation. The mechanism for this highly coordinated process is poorly understood. Here we show that dynasore (DNSR), a small-molecule inhibitor of dynamins (DNMs), or short hairpin RNA knockdown of DNM2 and DNM3 impairs directional migration in a human MK cell line or MKs derived from cultured CD34+ cells. Because cell migration requires actin cytoskeletal rearrangements, we measured actin polymerization and the activity of cytoskeleton regulator RhoA and found them to be decreased after inhibition of DNM2 and DNM3. Because SDF-1α is important for hematopoiesis, we studied the expression of its receptor CXCR4 in DNSR-treated cells. CXCR4 expression on the cell surface was increased, at least partially because of slower endocytosis and internalization after SDF-1α treatment. Combined inhibition of DNM2 and DNM3 or forced expression of dominant-negative Dnm2-K44A or GTPase-defective DNM3 diminished ß1 integrin (ITGB1) activity. DNSR-treated MKs showed an abnormally clustered staining pattern of Rab11, a marker of recycling endosomes. This suggests decreased recruitment of the recycling pathway in DNSR-treated cells. Altogether, we show that the GTPase activity of DNMs, which governs endocytosis and regulates cell receptor trafficking, exerts control on MK migration toward SDF-1α gradients, such as those originating from the vascular niche. DNMs play a critical role in MKs by triggering membrane-cytoskeleton rearrangements downstream of CXCR4 and integrins.

Herpesvirus-encoded microRNAs detected in human gingiva alter host cell transcriptome and regulate viral infection.

MicroRNAs (miRNAs) are small, non-coding RNAs of ~18-25 nucleotides that have gained extensive attention as critical regulators in complex gene networks including immune cell lineage commitment, differentiation, maturation, and maintenance of immune homeostasis and function. Many viruses encode miRNAs that directly downregulate the expression of factors of the innate immune system, which includes proteins involved in promoting apoptosis and recruitment. In this study, we examined the expression profiles of three previously identified viral miRNAs (v-miRs) from the human herpesvirus (HHV) family, HSV-1 (miR-H1), KSHV (miR-K12-3-3p), and HCMV (miR-US4) in healthy and diseased periodontal tissues and observed increased levels of v-miRs in diseased tissues. To understand the significance of this increase, we overexpressed v-miRs in human oral keratinocytes (HOK), a common target for various HHV, and analyzed the impact of miR-H1 and miR-K12-3-3p on the host transcriptome. More than 1300 genes were altered in HOK overexpressing miR-H1 and miR-K12-3-3p. Global pathway analysis of deregulated genes identified several key cellular pathways that may favor viral persistence. Using bioinformatic analysis, we predicted hundreds of potential v-miR binding sites on genes downregulated by miR-H1 and miR-K12-3-3p and validated three novel target v-miR sites suggesting widespread direct and indirect modulation of numerous host genes/pathways by a single v-miR. Finally, in vitro HSV-1 infection assays showed that miR-H1 can regulate viral entry and infection in human oral keratinocytes (HOK). Overall, our results demonstrate clinical and functional relevance of pathogenic viral molecules viz., v-miRs that regulate both host and viral functions and may contribute to the pathogenesis of inflammatory oral diseases.

Transcriptional Regulation of Emergency Granulopoiesis in Leukemia.

Neutropenic conditions are prevalent in leukemia patients and are often associated with increased susceptibility to infections. In fact, emergency granulopoiesis (EG), a process regulating neutrophil homeostasis in inflammatory conditions and infections, may occur improperly in leukemic conditions, leading to reduced neutrophil counts. Unfortunately, the mechanisms central to dysfunctional EG remain understudied in both leukemia patients and leukemic mouse models. However, despite no direct studies on EG response in leukemia are reported, recently certain transcription factors (TFs) have been found to function at the crossroads of leukemia and EG. In this review, we present an update on TFs that can potentially govern the fate of EG in leukemia. Transcriptional control of Fanconi DNA repair pathway genes is also highlighted, as well as the newly discovered role of Fanconi proteins in innate immune response and EG. Identifying the TFs regulating EG in leukemia and dissecting their underlying mechanisms may facilitate the discovery of therapeutic drugs for the treatment of neutropenia.

Terminal Maturation of Orthochromatic Erythroblasts Is Impaired in Burn Patients.

Mechanisms of erythropoietin (Epo)-resistant anemia in burn patients are poorly understood. We have recently found that administering a nonselective beta 1,2-adrenergic blocker propranolol (PR) was effective in reversing myelo-erythroid commitment through MafB regulation and increase megakaryocyte erythrocyte progenitors in burn patients' peripheral blood mononuclear cell (PBMC)-derived ex vivo culture system. Having known that Epo-dependent proliferation of early erythroblasts is intact after burn injury, here we inquired whether or not Epo-independent maturation stage of erythropoiesis is affected by burn injury and the relative role of PR on late-stage erythropoiesis. While majority of erythropoiesis occurs in the bone marrow, maturation into reticulocytes is crucial for their release into sinusoids to occupy the peripheral circulation for which enucleation is vital. peripheral blood mononuclear cells (PBMCs) from burn patients were extended beyond commitment and proliferation stages to late maturation stage in ex vivo culture to understand the role of PR in burn patients. Burn impedes late maturation of orthochromatic erythroblasts into reticulocytes by restricting the enucleation step. Late-stage erythropoiesis is impaired in burn patients irrespective of PR treatment. Further, substituting the microenvironment with control plasma (homologous) in place of autologous plasma rescues the conversion of orthochromatic erythroblasts to reticulocytes. Results show promise in formulating interventions to regulate late-stage erythropoiesis, which can be used in combination with PR to reduce the number of transfusions.

Brief Report: Radiofrequency Ablation Therapy for Anal Intraepithelial Neoplasia: Results From a Single-Center Prospective Pilot Study in HIV+ Participants.

BACKGROUND: HIV-positive individuals have high incidence of anal high-grade squamous intraepithelial lesions (HSIL) at the squamocolumnar junction (SCJ), which can progress to cancer. Focal radiofrequency ablation (RFA) treats HSIL, but metachronous recurrence remains high and may be improved with circumferential treatment. SETTING: The study was performed at a single center. METHODS: This was a prospective trial of circumferential anal RFA using Barrx 60 RFA focal catheters in participants with ≥1 anal SCJ HSIL. The entire SCJ was ablated by RFA. Adverse events (AEs), symptoms, including pain, and quality of life were assessed. High-resolution anoscopy assessed recurrences at 3, 6, 9, and 12 months. Lesion site biopsies occurred at month 12. Recurrences were retreated with focal RFA. RESULTS: Ten male participants (9 HIV+), with a median 2 HSILs (range 2-8) each, enrolled. Median T-cell count and viral load were 730 cells/mcL and 38 copies/mL. Median duration of RFA treatment was 6.5 (5-13) minutes. Lesion persistence occurred in 4 participants (3 at 3 months, 1 at 6 months). Recurrence at a new site occurred in 1 participant at 3 months. No lesion persisted after retreatment. All participants were HSIL free and completely healed by 12 months. Two device-related mild AEs occurred in 1 participant each (thrombosed external hemorrhoid and soft anal scar; both resolved). No serious AEs occurred. CONCLUSION: Circumferential anal SCJ RFA produced total HSIL eradication with no more than 2 treatments. Circumferential RFA seems to be well tolerated with minimal pain and no serious AEs in HIV+ participants. CLINICALTRIALS.GOV:: NCT02189161.

Exacerbation of N-nitrosodiethylamine Induced Hepatotoxicity and DNA Damage in Mice Exposed to Chronic Unpredictable Stress.

Psychological stress contributes to increased susceptibility to a number of diseases including cancer. The present study was designed to assess the effect of chronic unpredictable stress on N-nitrosodiethylamine induced liver toxicity in terms of in vivo antioxidant status and DNA damage in Swiss albino mice. The animals used in this study were randomized into different groups based on the treatment with N-nitrosodiethylamine or chronic unpredictable stress alone and post-stress administration of N-nitrosodiethylamine. The mice were sacrificed after 12 weeks of treatment, and the status of major enzymatic and non-enzymatic antioxidants, liver function markers, lipid peroxidation and the extent of DNA damage were determined in circulation and liver tissues of all the groups. The N-nitrosodiethylamine treated group showed significantly compromised levels of the antioxidant enzymes, lipid peroxidation, and the liver function markers with enhanced DNA damage as compared to chronic unpredictable stress or control groups. A similar but less typical pattern observed in the chronic unpredictable stress treated mice. All the measured biochemical parameters were significantly altered in the group treated with the combination of chronic unpredictable stress and N-nitrosodiethylamine when compared to controls, or chronic unpredictable stress alone and/or N-nitrosodiethylamine alone treated groups. Thus, exposure to continuous, unpredictable stress conditions even in general life may significantly enhance the hepatotoxic potential of N-nitrosodiethylamine through an increase in the oxidative stress and DNA damage.

Discrete ß-adrenergic mechanisms regulate early and late erythropoiesis in erythropoietin-resistant anemia.

BACKGROUND: Anemia of critical illness is resistant to exogenous erythropoietin. Packed red blood cells transfusions is the only treatment option, and despite related cost and morbidity, there is a need for alternate strategies. Erythrocyte development can be divided into erythropoietin-dependent and erythropoietin-independent stages. We have shown previously that erythropoietin-dependent development is intact in burn patients and the erythropoietin-independent early commitment stage, which is regulated by ß1/ß2-adrenergic mechanisms, is compromised. Utilizing the scald burn injury model, we studied erythropoietin-independent late maturation stages and the effect of ß1/ß2, ß-2, or ß-3 blockade in burn mediated erythropoietin-resistant anemia. METHODS: Burn mice were randomized to receive daily injections of propranolol (nonselective ß1/ß2 antagonist), nadolol (long-acting ß1/ß2 antagonist), butoxamine (selective ß2 antagonist), or SR59230A (selective ß3 antagonist) for 6 days after burn. Total bone marrow cells were characterized as nonerythroid cells, early and late erythroblasts, nucleated orthochromatic erythroblasts and enucleated reticulocyte subsets using CD71, Ter119, and Syto-16 by flow cytometry. Multipotential progenitors were probed for MafB expressing cells. RESULTS: Although propranolol improved early and late erythroblasts, only butoxamine and selective ß3-antagonist administrations were positively reflected in the peripheral blood hemoglobin and red blood cells count. While burn impeded early commitment and late maturation stages, ß1/ß2 antagonism increased the early erythroblasts through commitment stages via ß2 specific MafB regulation. ß3 antagonism was more effective in improving overall red blood cells through late maturation stages. CONCLUSION: The study unfolds novel ß2 and ß3 adrenergic mechanisms orchestrating erythropoietin resistant anemia after burn, which impedes both the early commitment stage and the late maturation stages, respectively.

Myelo-erythroid commitment after burn injury is under ß-adrenergic control via MafB regulation.

Severely injured burn patients receive multiple blood transfusions for anemia of critical illness despite the adverse consequences. One limiting factor to consider alternate treatment strategies is the lack of a reliable test platform to study molecular mechanisms of impaired erythropoiesis. This study illustrates how conditions resulting in a high catecholamine microenvironment such as burns can instigate myelo-erythroid reprioritization influenced by ß-adrenergic stimulation leading to anemia. In a mouse model of scald burn injury, we observed, along with a threefold increase in bone marrow LSK cells (linneg Sca1+cKit+), that the myeloid shift is accompanied with a significant reduction in megakaryocyte erythrocyte progenitors (MEPs). ß-Blocker administration (propranolol) for 6 days after burn, not only reduced the number of LSKs and MafB+ cells in multipotent progenitors, but also influenced myelo-erythroid bifurcation by increasing the MEPs and reducing the granulocyte monocyte progenitors in the bone marrow of burn mice. Furthermore, similar results were observed in burn patients' peripheral blood mononuclear cell-derived ex vivo culture system, demonstrating that commitment stage of erythropoiesis is impaired in burn patients and intervention with propranolol (nonselective ß1,2-adrenergic blocker) increases MEPs. Also, MafB+ cells that were significantly increased following standard burn care could be mitigated when propranolol was administered to burn patients, establishing the mechanistic regulation of erythroid commitment by myeloid regulatory transcription factor MafB. Overall, results demonstrate that ß-adrenergic blockers following burn injury can redirect the hematopoietic commitment toward erythroid lineage by lowering MafB expression in multipotent progenitors and be of potential therapeutic value to increase erythropoietin responsiveness in burn patients.

A trial of radiofrequency ablation for anal intraepithelial neoplasia.

PURPOSE: Radiofrequency ablation (RFA) effectively treats esophageal high-grade dysplasia, but its efficacy in treating anal canal high-grade squamous intraepithelial lesions (HSILs) is unsubstantiated. This prospective study assessed the safety and efficacy of applying hemi-circumferential RFA to anal canal HSIL. METHODS: Twenty-one HIV-negative participants with HSIL occupying ≤ half the anal canal circumference were treated with hemi-circumferential anal canal RFA. Participants were assessed every 3 months for 12 months with high-resolution anoscopy; recurrence in the treatment zone was re-treated with focal RFA. RESULTS: Twenty-one participants with a mean of 1.7 lesions (range 1-4) enrolled and completed the trial. Six (29 %) participants had recurrent HSIL within the treated hemi-circumference within 1 year. Four participants (19 %) had persistence of an index lesion at 3 months. One (2.9 %) index HSIL persisted again at 12 months. No participants had more than two RFA treatments. KM curve-predicted HSIL-free survival within the treatment zone at 1 year was 76 % (95 % CI 52-89 %). Comparing the first 7 and last 14 participants, the predicted 1-year HSIL-free survivals are 43 % (95 % CI 10-73 %) and 93 % (95 % CI 59-99 %), respectively (p = 0.008), suggesting a learning curve with the treating physician. Multivariable analysis showed decreased recurrence in the last 14 participants (HR 0.02; 95 % CI 0.001-0.63) while increasing BMI increased recurrence (HR 1.43, 95 % CI 1.01-2.01). No participants had device or procedure-related serious adverse events, anal stricture, or heavy bleeding. CONCLUSIONS: Hemi-circumferential RFA yielded a high rate of anal HSIL eradication in HIV-negative patients at 1 year with minimal adverse events. Lesion persistence was probably related to incomplete initial ablation.

Combined treatment of epigallocatechin gallate and Coenzyme Q10 attenuates cisplatin-induced nephrotoxicity via suppression of oxidative/nitrosative stress, inflammation and cellular damage.

Cisplatin (CP), a platinum based anticancer drug is used as one of the first-line therapy for the treatment of different types of solid tumors. However, CP-induced side effects particularly, nephrotoxicity is a major concern. A single nephrotoxic dose (7 mg/kg body weight) of CP was administered in rats with or without, pre and post combined multidoses of epigallocatechin gallate (EGCG) and coenzyme Q10 (CoQ10) (15 and 5 mg/kg body weight respectively). CP administration resulted in marked increase in the nephrotoxic parameters with alterations in the oxidative and nitrosative stress markers. The concentration of inflammatory, as well as apoptotic markers were markedly up-regulated in the kidney of the CP-treated group. Furthermore, CP resulted in histological injury in the renal tissues. Combined antioxidant treatment significantly (p < 0.01) attenuated CP-induced oxidative stress, nitrosative stress, inflammatory and apoptotic parameters. Moreover, an improvement in the histopathological changes confirmed the nephroprotective effect of antioxidant treatment. In conclusion, our study indicates that the combinatorial multidoses of EGCG and CoQ10 ameliorate the cisplatin-mediated pathogenesis by improving renal oxidative/nitrosative status, inflammation and apoptosis and thus can be used as a promising protective agent to increase the efficacy of the drug by minimizing its major side effect i.e. nephrotoxicity.