Early Versus Standard Initiation of Terlipressin for Acute Kidney Injury in ACLF: A Randomized Controlled Trial (eTerli Study).

BACKGROUND AND AIMS: Terlipressin infusion is effective in hepatorenal syndrome (HRS-AKI). However, its efficacy for HRS-AKI resolution in acute-on-chronic liver failure (ACLF) patients has been suboptimal. Progression of AKI is rapid in ACLF. We investigated whether early initiation of terlipressin(eTerli) can improve response rates. METHODS: Consecutive ACLF patients with stage II/III AKI despite albumin resuscitation (40 g) were randomized to receive terlipressin at 2 mg/24 h plus albumin at 12 h (ET, n = 35) or at 48 h as standard therapy (ST, n = 35). (June 22, 2020 to June 10, 2022). The primary end-point was AKI reversal by day7. RESULTS: Baseline parameters including AKI stage and ACLF-AARC scores in two arms were comparable. Full AKI response at day 7 was higher in ET [24/35 (68.6%)] than ST arm [11/35 (31.4%; P 0.03]. Day3 AKI response was also higher in ET arm [11/35 (31.4%) vs. 4/35 (11.4%), P 0.04]. Using ST compared to ET [HR 4.3; P 0.026] and day 3 serum creatinine > 1.6 mg/dl [HR 9.1; AUROC-0.866; P < 0.001] predicted HRS-AKI non-response at day 7. ET patients showed greater improvement in ACLF grade, mean arterial pressure, and urine output at day 3, and required lower albumin within 7 days than ET arm (149.1 ± 41.8 g vs. 177.5 ± 40.3 g, P 0.006) and had lower 28-day mortality: 40% vs. 65.7%, P 0.031]. Early use of terlipressin than ST [HR 2.079; P 0.038], baseline HE [HR 2.929; P 0.018], and AKI persistence at day 3 [HR 1.369; P 0.011] predicted 28-day mortality. Fifteen (21.4%) patients had treatment related adverse effects, none was life threatening. CONCLUSION: In ACLF patients, early initiation of terlipressin for AKI persisting after 12 h of volume expansion with albumin helps in reduced short-term mortality and early AKI reversal with regression of ACLF stage. These results indicate need for change in current practice for terlipressin usage in HRS-AKI.

Higher circulating natural killer cells and lower lactate levels at admission predict spontaneous survival in non-acetaminophen induced acute liver failure.

Massive cellular necrosis in acute liver failure (ALF) is dominantly immune mediated and innate immune cells are major pathophysiological determinants in liver damage. In fifty ALF and fifteen healthy, immune cells phenotyping by flow-cytometry, DAMPs using ELISA were analysed and correlated with clinical and biochemical parameters. ALF patients (aged 27 ± 9 yr, 56% males, 78% viral aetiology) showed no difference in neutrophils and classical monocytes, but significantly increased intermediate monocytes (CD14+CD16+) (p < 0.01), decreased non-classical monocytes (CD14-CD16+) and CD3-veCD16+CD56+ NK cells compared to HC. ALF patients who survived, showed higher NK cells (9.28 vs. 5.1%, p < 0.001) among lymphocytes and lower serum lactate levels (6.1 vs. 28, Odds ratio 2.23, CI 1.27-3.94) than non- survivors had higher. Logistic regression model predicted the combination of lactate levels with NK cell percentage at admission for survival. In conclusion, Combination of NK cell frequency among lymphocytes and lactate levels at admission can reliably predict survival of ALF patients.

Standardisation of neonatal clinical practice.

The International Fetal and Newborn Growth Consortium for the 21(st) Century (INTERGROWTH-21(st) ) is a large-scale, population-based, multicentre project involving health institutions from eight geographically diverse countries, which aims to assess fetal, newborn and preterm growth under optimal conditions. Given the multicentre nature of the project and the expected number of preterm births, it is vital that all centres follow the same standardised clinical care protocols to assess and manage preterm infants, so as to ensure maximum validity of the resulting standards as indicators of growth and nutrition with minimal confounding. Moreover, it is well known that evidence-based clinical practice guidelines can reduce the delivery of inappropriate care and support the introduction of new knowledge into clinical practice. The INTERGROWTH-21(st) Neonatal Group produced an operations manual, which reflects the consensus reached by members of the group regarding standardised definitions of neonatal morbidities and the minimum standards of care to be provided by all centres taking part in the project. The operational definitions and summary management protocols were developed by consensus through a Delphi process based on systematic reviews of relevant guidelines and management protocols by authoritative bodies. This paper describes the process of developing the Basic Neonatal Care Manual, as well as the morbidity definitions and standardised neonatal care protocols applied across all the INTERGROWTH-21(st) participating centres. Finally, thoughts about implementation strategies are presented.

Podoplanin-positive dilated lymphatic vessels in duodenum associates with three-month mortality in patients with cirrhosis.

Dilated and dysfunctional gut lymphatic vessels (LVs) have been reported in experimental cirrhosis. Here, we studied LVs in duodenal (D2)-biopsies of liver cirrhosis patients and investigated the prognostic role of a LV marker, podoplanin (PDPN), in predicting the mortality of patients with cirrhosis. A prospective, single-center cohort study was performed in liver cirrhosis patients (n = 31) and matched healthy controls (n = 9). D2-biopsies were obtained during endoscopy procedure, immunostained with PDPN, and scored based on 1) intensity and 2) density of positively-stained LVs per high power field. Gut and systemic inflammation were estimated by quantifying duodenal CD3+ intraepithelial lymphocytes (IELs), CD68+ macrophages, and serum TNF-α and IL-6 levels, respectively. Gut permeability and inflammation as assessed by quantifying gene expression of TJP1, OCLN, TNF-α, and IL-6 in D2-biopsies. Gene expression of LV markers, PDPN (8-fold), and LYVE1 (3-fold) was enhanced in D2-biopsies of cirrhosis patients compared to control (p < 0.0001). The mean PDPN score in decompensated cirrhosis patients (6.91 ± 1.26, p < 0.0001) was significantly increased as compared to those with compensated (3.25 ± 1.60). PDPN score positively and significantly correlated with the number of IELs (r = 0.33), serum TNF-α (r = 0.35), and IL-6 (r = 0.48) levels, while inversely correlated with TJP1 expression (r = -0.46, p < 0.05 each). In Cox regression, the PDPN score was a significant and independent 3-month-mortality predictor in patients (HR: 5.61; 1.08-29.109; p = 0.04). The area under the curve for the PDPN score was 84.2, and cutoff value for predicting mortality was ≥6.5 with 100% sensitivity and 75% specificity. Collectively, dilated LVs with high PDPN expression in D2-biopsies is a characteristic feature of patients with decompensated cirrhosis. PDPN score correlates with enhanced gut and systemic inflammation and also associates with 3-month mortality in cirrhosis.

The Role of Leadership in Public Sector Innovation: A Systematic Review and Meta-Analysis of the Management of COVID-19 in Asian Countries.

Strong leadership in public sector innovation can empower governments to address community challenges in new ways in light of the challenges posed by the global coronavirus pandemic. Coronavirus management policy, pandemic responses, needs, and options are reflected in various Asian countries in respective published literature, but a summarized synthesis is not available. Using a systematic review approach (PRISMA), this study has analyzed the role of leadership in public sector innovation in COVID-19 management and synthesized 23 articles from 23 different Asian countries. In the light of available data, public sector innovation (PSI) and the role played by the leadership of each country' have been found to be largely inter-dependent. The current review provides a cross-section of the ongoing nature of the pandemic, as management responses and trend data in the countries are still emerging or evolving. Additionally, our study contributes a current state report regarding the barriers facing the leadership of Asian countries in mitigating the global pandemic through PSI. Our study found that a strong political leadership presence combined with a technocratic approach and a highly-skilled public sector workforce, could lead to more tremendous success in managing the outbreak. Furthermore, religious leadership was also found to have a potentially significant role in COVID-19 management strategies.

Prevalence and the factors influencing soil-transmitted helminths among school age children (5-14 years age) in a rural area of Coimbatore district.

CONTEXT: Highest intensity of soil-transmitted helminthiasis (STH) is seen among school age children. AIMS: The aim of this study is to find out the prevalence and factors associated with soil-transmitted helminthic infection among school age children (5-14 years) in a rural area of Coimbatore district. SETTINGS AND DESIGN: The study was conducted in the field practice area of the Rural Health Training Centre (RHTC) Vedapatti, Coimbatore. RHTC caters to a total population of 23,841 distributed in 14 villages. After getting ethical clearance, five of the 14 villages of Vedapatti were selected by the cluster sampling method. Totally, 819 participated in the survey conducted between November 2015 and July 2016 in the field practice area. SUBJECTS AND METHODS: Structured questionnaire was used to collect the information. Consent from parents and assent from child were obtained. Totally, 610 gave one adequate stool sample. Early morning samples were collected and transported to the laboratory within four hours. Formal ether concentration method was performed, and examination was done. STATISTICAL ANALYSIS USED: Data analysis was performed with the SPSS version 19 software. The prevalence is expressed in percentage with 95% confidence interval (CI). Univariate and multivariate analyses were performed. Strength of association was expressed in terms of odds ratio (OR) and adjusted OR with 95% CI. P < 0.05 was considered as statistically significant. RESULTS: The prevalence of STH was 7.70% (95% CI: 5.58-9.82). Ascaris lumbricoides was highly prevalent 6.9% (4.89%-8.91%) followed by Hook worm 0.7% (0.04%-1.36%), and Trichuris trichura 0.2% (0.15%-0.55%). Mulitivariate logistic regression analysis showed that pucca houses offered protection against STH. CONCLUSIONS: The prevalence of STH in a rural area of Coimbatore is 7.7% (95% CI: 5.58-9.82), and is continuing as a public health problem.

Tracheostomy in critically ill liver disease patients with coagulopathy: A retrospective study at a tertiary center.

INTRODUCTION: Critically ill patients with liver disease commonly present to the intensive care unit (ICU) with need for prolonged ventilation, difficult weaning, and refractory coagulopathy. These patients experience both bleeding and thrombotic complications with a precariously balanced state of coagulopathy. The purpose of this study was to assess the bleeding complications of tracheostomy in critically ill patients with liver disease. METHODS: A retrospective study was conducted in liver ICU of a tertiary teaching institute. Medical records were analyzed to assess postprocedure complication rate among 73 critically ill liver disease patients who had undergone tracheostomy during the period of October 2017 to September 2018. RESULTS: Ten out of 73 patients (13%) required transfusion of blood products after 12 h of procedure, despite thromboelastography (TEG)-based correction prior to procedure. Of these, 7 patients (9%) underwent surgical tracheostomy (ST) and three patients (4%) underwent percutaneous tracheostomy. Statistically no significant difference in bleeding was seen among the two groups, but a rising trend was seen with the ST group (P = 0.52). None of the patients experienced procedure-related pneumothorax and subcutaneous emphysema, as observed in the chest X-ray. CONCLUSION: We conclude that coagulopathy should not be deterrence for the performance of tracheostomy in critically ill patients with liver disease. Adequate clotting support guided by the global tests of coagulation, such as TEG, ensures lesser incidence of bleeding.

When Mount Fuji Can Erupt after Seven Days: A Case Report of Delayed Posttraumatic Tension Pneumocephalus with Literature Review.

Tension pneumocephalus (TPC) is a neurosurgical emergency that occurs when there is an expansion of trapped intracranial gas causing raised intracranial pressure. Rarely, posttraumatic TPC can occur even after 72 hours although the initial scans are normal. There are less than 20 cases of delayed TPC in the reported literature. Here, we report a case of delayed TPC that occurred 7 days after the initial injury and presented as sudden neurological deterioration. It was promptly diagnosed with a computed tomography brain and appropriate surgical intervention was performed and the outcome was good. We also did a literature review of reported cases of delayed TPC and looked out for factors that may predict its occurrence. The occurrence of an episode of cerebrospinal fluid rhinorrhea, followed by worsening of headache and sensorium in a patient with anterior cranial fossa fracture should alert a neurosurgeon to the possibility of delayed TPC.

Primary and secondary bystander effect and genomic instability in cells exposed to high and low linear energy transfer radiations.

Purpose: Non-Targeted effects (NTE), such as bystander effect (BE) and genomic instability (GI) challenge central dogma of radiation biology. Moreover, there is a need to understand its universality in different type of cells and radiation quality.Materials and method: To study BE (primary and secondary) and GI Human adult dermal fibroblast (HADF) and peripheral blood lymphocytes (PBL) were exposed to low fluence of 241Am alpha (α) particle and 6 MV X-ray. The BE was carried out by means of co-culture methodology after exposing the cells to both types of radiation and damage was measured using micronucleus assay (MN) and chromosomal aberration assay (CA) in the p1 cells while the GI was followed up in their progeny.Results: A dose-dependent increase in DNA damages (MN and CA) was observed in directly irradiated and bystander cells. The magnitude of BE was higher (6 fold) in cells co-cultured with the α-irradiated cells than that of with X-irradiated cells. Cross exposure of both cell types confirms that radiation induced BE is cell type dependent. In addition, induced DNA damage persisted for a longer population doubling in α-particle irradiated cells.Conclusion: This work adds evidence to secondary bystander response generated from primary bystander normal cells and its dependence to radiation quality.

Synthesis, characterisation and biomedical applications of curcumin conjugated chitosan microspheres.

Curcumin is a diaryl heptanoid of curcuminoids class obtained from Curcuma longa. It possesses various biological activities like anti-inflammatory, hypoglycemic, antioxidant, wound-healing, and antimicrobial activities. Chitosan is a biocompatible, biodegradable and non-toxic natural polymer which enhances the adhesive property of the skin. Chemical conjugation will leads to sustained release action and to enhance the bioavailability. This study aims to synthesis and characterize biocompatible curcumin conjugated chitosan microspheres for bio-medical applications. The Schiff base reaction was carried out for the preparation of curcumin conjugated chitosan by microwave method and it was characterised using FTIR and NMR. Curcumin conjugated chitosan microspheres (CCCMs) were prepared by wet milling solvent evaporation method. SEM analysis showed these CCCMs were 2-5µm spherical particles. The antibacterial activities of the prepared CCCMs were studied against Staphylococcus aureus and Escherichia coli, the zone of inhibition was 28mm and 23mm respectively. Antioxidant activity of the prepared CCCMs was also studied by DPPH and H2O2 method it showed IC50 esteem value of 216µg/ml and 228µg/ml, and anti-inflammatory activity results showed that CCCMs having IC50 value of 45µg/ml. The results conclude that the CCCMs having a good antibacterial, antioxidant and anti-inflammatory activities. This, the prepared CCCMs have potential application in preventing skin infections.

Carboxymethylated É©-carrageenan conjugated amphotericin B loaded gelatin nanoparticles for treating intracellular Candida glabrata infections.

Intercellular Candida glabrata infections are difficult to treat due to poor penetration of drugs into the fungal niche. Delivering amphotericin B (Amp B) into the macrophages where the pathogen inhabits is an effective solution. We are studying the macrophage targeting proficiency of É©-carrageenan for the delivery of Amp B using gelatin A nanoparticles (GNPs). The choice of gelatin A was the outcome of in silico inspections where the amino functionalized polymer having the best docking score with Amp B was selected. We prepared a sustained release formulation of amp B loaded carboxymethyl É©-carrageenan conjugated gelatin nanoparticles (CMC-Amp B-GNPs) with size 343±12nm and -25±5.3mV zeta potential. The formulations were found to be stable, biocompatible and non-haemolytic. Flow cytometry analysis showed 3 fold higher uptake of CMC-GNPs compared to the GNPs by RAW 264.7 cells. CMC-Amp B-GNPs showed enhanced antifungal activity than bare Amp B and Amp B-GNPs.

Quantification of optical Doppler broadening and optical path lengths of multiply scattered light by phase modulated low coherence interferometry.

We show experimental validation of a novel technique to measure optical path length distributions and path length resolved Doppler broadening in turbid media for different reduced scattering coefficients and anisotropies. The technique involves a phase modulated low coherence Mach-Zehnder interferometer, with separate fibers for illumination and detection. Water suspensions of Polystyrene microspheres with high scattering and low absorption levels are used as calibrated scattering phantoms. The path length dependent diffusion broadening or Doppler broadening of scattered light is shown to agree with Diffusive Wave Spectroscopy within 5%. The optical path lengths are determined experimentally from the zero order moment of the phase modulation peak around the modulation frequency in the power spectrum and the results are validated with Monte Carlo simulations.

Effect of speckles on the depth sensitivity of laser Doppler perfusion imaging.

A theoretical model is presented and experimentally validated that allows the prediction of the effect of speckles on the depth sensitivity of laser Doppler perfusion imaging. It is shown that the influence of speckles on depth sensitivity is large. In particular the sensitivity to particle motion in superficial layers is strongly beam diameter dependent: decreasing the beam diameter on the tissue surface increases the sensitivity to superficial motion to a much stronger extent than sensitivity to motion at a larger depth. This can be explained through the effect of beam diameter changes on the fractional coherence areas generated by photons with different penetration depths in the tissue.

Discrimination between Doppler-shifted and non-shifted light in coherence domain path length resolved measurements of multiply scattered light.

We show a novel technique to distinguish between Doppler shifted and unshifted light in multiple scattering experiments on mixed static and dynamic media. With a phase modulated low coherence Mach- Zehnder interferometer, optical path lengths of shifted and unshifted light and path length dependent Doppler broadening are measured in a two-layer tissue phantom, with a superficial static layer of different thickness covering a semi-infinite dynamic medium having identical optical properties. No Doppler broadening is observed until a certain optical path length depending on the thickness of the superficial static layer. From the minimum optical path length corresponding to the Doppler-shifted light the thickness of the static layer that overlies the dynamic layer can be estimated. Validation of the experimentally determined thickness of the static layer is done with the Doppler Monte Carlo technique. This approach has potential applications in discriminating between statically and dynamically scattered light in the perfusion signal and in determining superficial burn depths.

Controlled Delivery of Bioactive Molecules for the Treatment of Chronic Wounds.

A cut or break on the surface of the skin is usually referred to as a wound. Any wound has a potential to heal by itself through a complex cascade of events. However, some wounds show delayed healing due to their underlying physiology and are referred to as chronic wounds like diabetic ulcers, venous ulcers, pressure ulcers and chronic infected ulcers. Extensive care has to be taken for the management of chronic wounds and these have become a major concern in the current medical scenario. The use of bioactive molecules or in other words the molecules that can actively interact with the wound environment and help in wound healing are gaining much importance. The incorporation of bioactive molecules into a suitable matrix system which not only provide a controlled release of the molecules, but also enable better exudate management is desired to overcome the shortcomings of the conventional treatment modalities. A major problem associated with chronic wounds is that they are easily prone to infections. In such cases, the topical delivery of antibiotics helps eliminate infection. However, the continuous use of high dose of antibiotics has led to the development of multi drug resistant bacterial strains. To overcome these issues, other broad-spectrum antimicrobial agents like antiseptics, metallic nanoparticles and antimicrobial peptides are being adopted nowadays. Growth factors play a major role in the wound healing cascade, thus topical delivery of growth factor from a suitable matrix is an interesting strategy. The delivery of nucleic acids with the aid of suitable vectors for either silencing a particular gene or over expressing a gene of interest is also being investigated nowadays. This review is an attempt to draw light over some of the recent approaches adopted for the treatment of chronic wounds using bioactive molecules like antibiotics, antiseptics, metallic nanoparticles or ions, growth factors and nucleic acids.

Accelerated platelet activation in Asian Indians with diabetes and coronary artery disease--The Chennai Urban Population Study (CUPS-13).

AIM: To assess platelet activation in south Indian type 2 diabetic subjects with and without CAD. METHODS: Four groups of subjects were studied; Group 1 comprised of non-diabetic subjects without coronary artery disease (CAD) (n = 30). Type 2 diabetic subjects without CAD formed Group 2 (n = 30); Group 3 comprised of type 2 diabetic subjects with CAD (n = 30) and Group 4 consisted of non- diabetic subjects with CAD (n=14). CAD was diagnosed based on coronary angiographic evidence of severe double or triple vessel disease. Platelet activation was tested after an overnight fast in blood obtained from a bleeding wound at 1 minute post-incision (wound-induced activation) as well as venous blood stimulated in vitro with collagen, using whole blood flow cytometry. In subjects with CAD, aspirin was withdrawn for 7 days and nitrates for 24 hours. RESULTS: Collagen induced GP IIb/IIIa binding was significantly higher among diabetic subjects with (28.10 +/-19.89; p<0.05) and without CAD (21.02+/-19.62; p<0.05) and non-diabetic subjects with CAD (23.89+/-15.65; p<0.05) compared to non-diabetic subjects without CAD (11.69+/-13.69). Regression analysis showed collagen induced GP IIb/IIIa binding to be significantly associated with CAD [odds ratio (OR): 1.029, p = 0.025] and diabetes (OR: 1.037, p = 0.007). CONCLUSION: Increased platelet activation is seen in urban south Indians with diabetes and CAD.

Metabolism of high-density lipoproteins in cultured rat luteal cells.

The uptake of cholesterol from high-density lipoproteins (HDL) labeled with 125I and [3H]cholesterol was examined in cultured rat luteal cells. Luteal cells were incubated with labeled HDL, following which the metabolic fate of the apolipoproteins and cholesterol moieties of the receptor-bound HDL were examined. About 50% of the originally bound HDL apolipoproteins were released into the medium in 24 h by a temperature-dependent process while only 5% of the HDL cholesterol was released unmetabolized. Inclusion of unlabeled HDL in the chase incubation resulted in increased release of apolipoprotein-derived radioactive products without significant change in the release of unmetabolized cholesterol. 60% of the apolipoprotein-derived radioactivity could be precipitated with trichloroacetic acid; the remaining trichloroacetic acid-soluble radioactive fraction was identified as [125I]iodotyrosine. Gel filtration chromatography of the chase-released material showed that the trichloroacetic acid-precipitable products, which contained no detectable amounts of cholesterol, eluted over a range of molecular sizes (9-80 kDa). No intact HDL was retroendocytosed. About 80% of trichloroacetic acid-precipitable products could be immunoadsorbed on anti-apolipoprotein A-I antibody immobilized on CNBr-activated Sepharose, suggesting the presence of fragments containing apolipoprotein A-I. This material was also capable of reassociating with native HDL. Lysosomal inhibitors were partially effective in inhibiting the amount of trichloroacetic acid-soluble products formed. The lysosomal degradation appeared to have no role in the uptake of HDL-derived cholesterol. These studies demonstrate preferential and total uptake of HDL cholesterol by luteal cells, with concomitant degradation of the lipoprotein.

Differential uptake and metabolism of free and esterified cholesterol from high-density lipoproteins in the ovary.

Rat luteal cells utilize high-density lipoproteins (HDL) as a source of cholesterol for steroid synthesis. Both the free and esterified cholesterol of HDL are utilized by these cells. In this report, we have examined the relative uptake of free and esterified cholesterol of HDL by cultured rat luteal cells. Incubation of the cells with HDL labeled with [3H]cholesterol or [3H]cholesteryl linoleate resulted in 4-6-fold greater uptake of the free cholesterol compared to esterified cholesterol. The increased uptake of free cholesterol correlated with its utilization for progestin synthesis: utilization of HDL-derived free cholesterol was 3-6-fold higher than would be expected from its concentration in HDL. The differential uptake and utilization of free and esterified cholesterol was further examined using egg phosphatidylcholine liposomes containing cholesterol or cholesteryl linoleate as a probe. Liposomes containing free cholesterol were able to deliver cholesterol to luteal cells and support steroid synthesis in the absence of apolipoproteins, and the addition of apolipoprotein A-I (apo A-I) moderately increased the uptake and steroidogenesis. Similar experiments using cholesteryl linoleate/egg phosphatidylcholine liposomes showed that inclusion of apo A-I resulted in a pronounced increase in the uptake of cholesteryl linoleate and progestin synthesis. These experiments suggest that free cholesterol from HDL may be taken up by receptor-dependent and receptor-independent processes, whereas esterified cholesterol uptake requires a receptor-dependent process mediated by apolipoproteins.

Epigenetic therapy restores normal hematopoiesis in a zebrafish model of NUP98-HOXA9-induced myeloid disease.

Acute myeloid leukemia (AML) occurs when multiple genetic aberrations alter white blood cell development, leading to hyperproliferation and arrest of cell differentiation. Pertinent animal models link in vitro studies with the use of new agents in clinical trials. We generated a transgenic zebrafish expressing human NUP98-HOXA9 (NHA9), a fusion oncogene found in high-risk AML. Embryos developed a preleukemic state with anemia and myeloid cell expansion, and adult fish developed a myeloproliferative neoplasm (MPN). We leveraged this model to show that NHA9 increases the number of hematopoietic stem cells, and that oncogenic function of NHA9 depends on downstream activation of meis1, the PTGS/COX pathway and genome hypermethylation through the DNA methyltransferase, dnmt1. We restored normal hematopoiesis in NHA9 embryos with knockdown of meis1 or dnmt1, as well as pharmacologic treatment with DNA (cytosine-5)-methyltransferase (DNMT) inhibitors or cyclo-oxygenase (COX) inhibitors. DNMT inhibitors reduced genome methylation to near normal levels. Strikingly, we discovered synergy when we combined sub-monotherapeutic doses of a histone deacetylase inhibitor plus either a DNMT inhibitor or COX inhibitor to block the effects of NHA9 on zebrafish blood development. Our work proposes novel drug targets in NHA9-induced myeloid disease, and suggests rational therapies by combining minimal doses of known bioactive compounds.

Involvement of microtubules in lipoprotein degradation and utilization for steroidogenesis in cultured rat luteal cells.

Cells isolated from superovulated rat ovaries metabolize low density lipoprotein (LDL) and high density lipoprotein (HDL) of human or rat origin and use the lipoprotein-derived cholesterol as a precursor for progesterone production. Under in vitro conditions, both lipoproteins are internalized and degraded in the lysosomes, although degradation of HDL is of lower magnitude than that of LDL. In this report we have examined the role of cellular microtubules in the internalization and degradation of human LDL and HDL in cultured rat luteal cells. The microtubule depolymerizing agents colchicine, podophyllotoxin, vinblastine, and nocodazole as well as taxol, deuterium oxide, and dimethyl sulfoxide, which are known to rapidly polymerize cellular tubulin into microtubules, were used to block the function of microtubules. When these antimicrotubule agents were included in the incubations, degradation of the apolipoproteins of [125I]iodo-LDL and [125I]iodo-HDL by the luteal cells was inhibited by 50-85% compared to untreated control values. Maximum inhibitory effects were observed when the cells were preincubated with the inhibitor for at least 4 h at 37 C before treatment with the labeled lipoprotein. Lipoprotein-stimulated progesterone production by luteal cells was also inhibited by 50% or more in the presence of antimicrotubule agents. However, basal and hCG-stimulated progesterone production were unaffected by these inhibitors. The binding of [125I]iodo-LDL and [125I]iodo-HDL to luteal cell plasma membrane receptors was not affected by the microtubule inhibitors. Although binding was unaffected and degradation was impaired in the presence of the inhibitors, there was no detectable accumulation of undegraded lipoprotein within the cells during the 24 h of study. From this study we conclude that the uptake and utilization of LDL and HDL by cultured rat luteal cells are mediated by cellular microtubules.