Compatibility studies of acyclovir and lactose in physical mixtures and commercial tablets

This study documents drug-excipient incompatibility studies of acyclovir in physical mixtures with lactose and in different tablet brands. Differential scanning calorimetry (DSC) was initially used to assess compatibility of mixtures. The Fourier-transform infrared (FTIR) spectrum was also compared with the spectra of pure drug and excipient. Although DSC results indicated incompatibility with lactose, FTIR spectra were mostly unmodified due to overlapping peaks. Samples of isothermally stressed physical mixture were stored at 95 °C for 24 h. The residual drug was monitored using a validated high-performance liquid chromatography (HPLC) assay and data fitting to solid-state kinetic models was performed. The drug loss kinetics followed a diffusion model. The aqueous mixture of drug and excipient was heated in order to prepare an adduct mixture. HPLC analysis revealed one extra peak that was fractionated and subsequently injected into the liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) system. The MRM (Multiple Reaction Monitoring) chromatograms characterized the peak with molecular mass corresponding to an acyclovir-lactose Maillard reaction product. The presence of lactose in commercial tablets was checked using a new TLC method. Overall, the incompatibility of acyclovir with lactose was successfully evaluated using a combination of thermal methods and LC-MS/MS.     

The latency-associated nuclear antigen of Kaposi sarcoma-associated herpesvirus induces B cell hyperplasia and lymphoma

Kaposi sarcoma-associated herpesvirus (KSHV) is a human lymphotropic herpesvirus. It is implicated in B cell neoplasias such as primary effusion lymphoma and multicentric Castleman disease in AIDS patients. The KSHV latency-associated nuclear antigen (LANA) is consistently expressed in all KSHV-associated tumor cells and was shown to bind the tumor suppressor proteins p53 and pRb. To test LANA's contribution to lymphomagenesis in vivo we generated transgenic mice expressing LANA under the control of its own promoter, which is B cell specific. All of the transgenic mice developed splenic follicular hyperplasia due to an expansion of IgM+ IgD+ B cells and showed increased germinal center formation. We also observed lymphomas, implying that LANA can activate B cells and provide the first step toward lymphomagenesis.     

Interaction of CpG-oligodeoxynucleotides with Toll like receptor 9 induces apoptosis and modulates metaloproteinase-2 activity in human intestinal epithelium

Recent reports have indicated different effects of immunostimulatory sequences containing CpG-Oligodeoxynucleotides (ODN) on various immune cells. However, the exact role of CpG-ODN in the human gut is unclear. In the present study, we assessed potential effects of CpG-ODN on non lymphoid cell (intestinal epithelial cell line HT-29) on a dose-response and time-course basis. Intestinal epithelial cell line HT-29 was treated with CpG-ODN (CpG 2006) and lipopolysaccharide (LPS) at 5, 10, 25, 50 microg/ ml and 1, 5, 10 microg/ ml concentrations, respectively. Following treatments, dose- response and time-course cytotoxicity using a colorimetric method, Metaloproteinase-2 (MMP-2) activity (using gelatin zymography) and apoptosis (using annexin-v flowcytometry method) assays were performed. Chloroquine treatment was also used for its inhibitory effect on endosomal acidification process to verify specific CpG-ODN and Toll Like Receptor 9 (TLR9) interactions. Cytotoxicity analysis of CpG-ODN showed that CpG-ODN increased significantly the proliferation of CpG-ODN treated cells, as compared to untreated cells, at concentrations of 10-25 microg/ml (p < 0.05). Overall MMP-2 activity analysis showed significant differences between treated and untreated cells. However, minimal changes were observed when MMP-2 activity was assessed per cell. Moreover, CpG-ODN treated cells demonstrated an increasing apoptosis rate of 0.8 %, 6.46 % and 14.21% at concentrations of 5, 10, 25 microg/ml, respectively. Collectively, our data indicated that intestinal epithelial cell line HT-29 is highly responsive to CpG effect in vitro and exhibits modified activities. The direct CpG-ODN and TLR-9 interactions in HT-29 cells could provide new approaches in malignant tumor therapeutic strategies.     

TGFbeta1 induces mast cell apoptosis

Mast cells are potent effectors of the inflammatory response, playing an important role in atopy, bacterial immunity, and animal models of arthritis, multiple sclerosis, and heart disease. Hence controlling mast cell numbers and responsiveness is essential for preventing inflammatory disease. We demonstrate that the cytokine transforming growth factor (TGF) beta1 is a potent inducer of mast cell apoptosis, a finding that was consistent in cultured mouse bone marrow-derived mast cells, peritoneal mast cells, and human mast cells. Cell death appeared to be caused by TGF-mediated repression of interleukin-3 (IL-3) receptor expression and function, leading to mitochondrial damage and activation of an apoptotic cascade acting via p53 and caspases. Although IL-3 receptor expression was reduced within 1 day of TGFbeta1 stimulation, apoptosis required at least 3 days to occur. This delay in onset is postulated to allow protective mast cell effector functions, protecting the host from infection while preventing the establishment of chronic inflammation. Our data support the theory that TGFbeta1 is an inhibitor of mast cell survival. The widespread expression of TGFbeta1 offers this cytokine as an ideal candidate for control of mast cell homeostasis.     

Tailored risk assessment of 90‐day acute heart failure readmission or all‐cause death to heart failure with preserved versus reduced ejection fraction

BackgroundAfter incident heart failure (HF) admission, patients are vulnerable to readmission or death in the 90‐day post‐discharge. Although risk models for readmission or death incorporate ejection fraction (EF), patients with HF with preserved EF (HFpEF) and those with HF with reduced EF (HFrEF) represent distinct cohorts. To better assess risk, this study developed machine learning models and identified risk factors for the 90‐day acute HF readmission or death by HF subtype.Methods and ResultsApproximately 1965 patients with HFpEF and 1124 with HFrEF underwent an index admission. Acute HF rehospitalization or death occurred in 23% of HFpEF and 28% of HFrEF groups. Of the 101 variables considered, multistep variable selection identified 24 and 25 significant factors associated with 90‐day events in HFpEF and HFrEF, respectively. In addition to risk factors common to both groups, factors unique to HFpEF patients included cognitive dysfunction, low‐pulse pressure, β‐blocker, and diuretic use, and right ventricular dysfunction. In contrast, factors unique to HFrEF patients included a history of arrhythmia, acute HF on presentation, and echocardiographic characteristics like left atrial dilatation or elevated mitral E/A ratio. Furthermore, the model tailored to HFpEF (area under the curve [AUC] = 0.770; 95% confidence interval [CI] 0.767–0.774) outperformed a model for the combined groups (AUC = 0.759; 95% CI 0.756–0.763).ConclusionThe UF 90‐day post‐discharge acute HF Re admission or Death Risk Assessment (UF90‐RADRA) models help identify HFpEF and HFrEF patients at higher risk who may require proactive outpatient management.     

Saving space: Comparing mini - logMAR with standard logMAR visual acuity

Purpose:Assessing visual acuity (VA) is the cornerstone of an ophthalmic workup and needs VA charts in a four or six meters space. The objective of this study was to compare the performance of distant VA (DVA) on one meter mini-logMAR (MLM) with a standard six meter logMAR (SLM) chart.Methods:We developed a MLM chart to be used at 1 m with +1.0 D spectacles, by reducing the SLM chart designed for 6 meters, to 1/6^th its size, using AutoCAD version 2014. On an initial cohort, we obtained DVA on the two charts by optometrist trainees, masked to the outcomes on the different tests. We performed regression and checked agreement between the two measurements. Subsequently, on a new cohort, we validated the performance of the MLM.Results:Of the 56 initial subjects, mean DVA with SLM was 0.44 ± 0.13 and with MLM was 0.45 ± 0.13; mean difference of -0.01 ± 0.02, 95%CI: 0.007 to 0.018; P < 0.0001 on paired t-test. There was a significant correlation: r = 0.99; r² = 0.98, P < 0.0001. On an average, DVA with MLM was less than a letter worse than with SLM. The regression formula obtained: SLM DVA = -0.1312 + 1.0014 x (MLM DVA). The validation study revealed no significant difference (P = 0.29) between the predicted standard DVA calculated by the regression formula and the actual standard.Conclusion:We suggest that we can deduce distance logMAR VA from a mini-logMAR chart as devised and used by us. This will take less space, be portable and allow congenial interaction with patients.     

Gene expression comparison between primary estrogen receptor-positive and triple-negative breast cancer with paired axillary lymph node metastasis

The aim of this study is to characterize and compare changes in gene expression patterns of paired axillary lymph node (ALN) metastases from estrogen receptor (ER)-positive and triple-negative (TNBC) primary breast cancer (PBC). Patients with stage 2-3 PBC with macrometastasis to an ALN were selected. Gene expression of 2567 cancer-associated genes was analyzed with the HTG EdgeSeq system coupled with the Illumina Next Generation Sequencing (NGS) platform. Changes in gene expression between ER/PR-positive, HER2-negative PBC, and their paired ALN metastases were compared with TNBC and their paired ALN metastases. Fourteen pairs of ER-positive and paired ALN metastasis were analyzed. Compared with the PBC, ALN metastasis had 673 significant differentially expressed genes, including 348 upregulated genes and 325 downregulated genes. Seventeen pairs of TNBC and paired ALN metastasis were analyzed. ALN metastasis had 257 significant differentially expressed genes, including 123 upregulated genes and 134 downregulated genes. When gene expression of the ALN for ER-positive PBC was compared to that of TNBC, 97 genes were upregulated in both, and 115 genes were similarly downregulated. Common upregulated genes were associated with cell death, necrosis, and homeostasis. Common downregulated genes were those of migration, degradation of extracellular matrix, and invasion. Although ER-positive PBC and TNBC have a distinct gene expression profiles and distinct changes from PBC to ALN metastases, a significant number of genes are similarly up- or downregulated. Understanding the role of these common genomic changes may provide clues to understanding the metastatic process itself.