We previously reported that autologous-fecal-microbiota-transplantation (aFMT), following 6 m of lifestyle intervention, attenuated subsequent weight regain and insulin rebound for participants ...consuming a high-polyphenol green-Mediterranean diet. Here, we explored whether specific changes in the core (abundant) vs. non-core (low-abundance) gut microbiome taxa fractions during the weight-loss phase (0-6 m) were differentially associated with weight maintenance following aFMT. Eighty-two abdominally obese/dyslipidemic participants (age = 52 years; 6 m weightloss = −8.3 kg) who provided fecal samples (0 m, 6 m) were included. Frozen 6 m's fecal samples were processed into 1 g, opaque and odorless aFMT capsules. Participants were randomly assigned to receive 100 capsules containing their own fecal microbiota or placebo over 8 m-14 m in ten administrations (adherence rate > 90%). Gut microbiome composition was evaluated using shotgun metagenomic sequencing. Non-core taxa were defined as ≤ 66% prevalence across participants. Overall, 450 species were analyzed. At baseline, 13.3% were classified as core, and Firmicutes presented the highest core proportion by phylum. During 6 m weight-loss phase, abundance of non-core species changed more than core species (P < .0001). Subject-specific changes in core and non-core taxa fractions were strongly correlated (Jaccard Index; r = 0.54; P < .001). Following aFMT treatment, only participants with a low 6 m change in core taxa, and a high change in non-core taxa, avoided 8-14 m weight regain (aFMT = −0.58 ± 2.4 kg, corresponding placebo group = 3.18 ± 3.5 kg; P = .02). In a linear regression model, low core/high non-core 6 m change was the only combination that was significantly associated with attenuated 8-14 m weight regain (P = .038; P = .002 for taxa patterns/treatment intervention interaction). High change in non-core, low-abundance taxa during weight-loss might mediate aFMT treatment success for weight loss maintenance.
ClinicalTrials.gov: NCT03020186
This study reports a highly efficient, rapid one-step purification process for the production of the recombinant vesicular stomatitis virus-based vaccine, rVSV-∆G-spike (rVSV-S), recently developed ...by the Israel Institute for Biological Research (IIBR) for the prevention of COVID-19. Several purification strategies are evaluated using a variety of chromatography methods, including membrane adsorbers and packed-bed ion-exchange chromatography. Cell harvest is initially treated with endonuclease, clarified, and further concentrated by ultrafiltration before chromatography purification. The use of anion-exchange chromatography in all forms results in strong binding of the virus to the media, necessitating a high salt concentration for elution. The large virus and spike protein binds very strongly to the high surface area of the membrane adsorbents, resulting in poor virus recovery (<15%), while the use of packed-bed chromatography, where the surface area is smaller, achieves better recovery (up to 33%). Finally, a highly efficient chromatography purification process with CaptoTM Core 700 resin, which does not require binding and the elution of the virus, is described. rVSV-S cannot enter the inner pores of the resin and is collected in the flow-through eluent. Purification of the rVSV-S virus with CaptoTM Core 700 resulted in viral infectivity above 85% for this step, with the efficient removal of host cell proteins, consistent with regulatory requirements. Similar results were obtained without an initial ultrafiltration step.
Apoptosis operates to eliminate damaged or potentially dangerous cells. This loss is often compensated by extra proliferation of neighboring cells. Studies in Drosophila imaginal discs suggest that ...the signal for the additional growth emanates from the dying cells. In particular, it was suggested that the initiator caspase Dronc mediates compensatory proliferation (CP) through Dp53 in wing discs. However, the exact mechanism that governs this CP remained poorly understood. We have previously shown that elimination of misspecified cells due to reduced Dpp signaling is achieved by the interaction of the co-repressor NAB with the transcriptional repressor Brk, which in turn induces Jun N-terminal kinase-dependent apoptosis. Here, we performed a systematic in vivo loss- and gain-of-function analysis to study NAB-induced death and CP. Our findings indicate that the NAB primary signal activates JNK, which in turn transmits two independent signals. One triggers apoptosis through the pro-apoptotic proteins Reaper and Hid, which in turn promote activation of caspases by the apoptosome components Ark and Dronc. The other signal induces CP in a manner that is independent of the death signal, Dronc, or Dp53. Once induced, the apoptotic pathway further activates a CP response. Our data suggest that JNK is the candidate factor that differentiates between apoptosis that involves CP and apoptosis that does not.
The antimicrobial peptide LL-37 belongs to the cathelicidin family and is the first amphipathic alpha-helical peptide isolated from human. LL-37 is considered to play an important role in the first ...line of defence against local infection and systemic invasion of pathogens at sites of inflammation and wounds. Understanding its mode of action may assist in the development of antimicrobial agents mimicking those of the human immune system. In vitro studies revealed that LL-37 is cytotoxic to both bacterial and normal eukaryotic cells. To gain insight into the mechanism of its non-cell-selective cytotoxicity, we synthesized and structurally and functionally characterized LL-37, its N-terminal truncated form FF-33, and their fluorescent derivatives (which retained structure and activity). The results showed several differences, between LL-37 and other native antimicrobial peptides, that may shed light on its in vivo activities. Most interestingly, LL-37 exists in equilibrium between monomers and oligomers in solution at very low concentrations. Also, it is significantly resistant to proteolytic degradation in solution, and when bound to both zwitterionic (mimicking mammalian membranes) and negatively charged membranes (mimicking bacterial membranes). The results also showed a role for the N-terminus in proteolytic resistance and haemolytic activity, but not in antimicrobial activity. The LL-37 mode of action with negatively charged membranes suggests a detergent-like effect via a 'carpet-like' mechanism. However, the ability of LL-37 to oligomerize in zwitterionic membranes might suggest the formation of a transmembrane pore in normal eukaryotic cells. To examine this possibility we used polarized attenuated total reflectance Fourier-transform infrared spectroscopy and found that the peptide is predominantly alpha-helical and oriented nearly parallel with the surface of zwitterionic-lipid membranes. This result does not support the channel-forming hypothesis, but rather it supports the detergent-like effect.
Summery Background Laparoscopic adjustable gastric banding (LAGB) surgery is the safest yet least effective method for massive weight loss. Thirty to 50 percent of patients undergoing LAGB will ...regain part of their lost weight. Objective To evaluate the effect of body contouring plastic surgery (BCPS) following LAGB on long-term body mass index (BMI) control. Setting Department of Surgery A, Soroka University Medical Center, Beer-Sheva, Israel. Methods A retrospective cohort study of patients aged 18-50 years, who underwent LAGB surgery between 1997 and 2007, was performed. Out of 2405 patients undergoing LAGB during those years, 861 were excluded and 1544 were recruited by phone. The final group included 72 patients matched for age and gender. Long-term weight regain was evaluated and compared between a group of patients who underwent subsequent body contouring plastic surgery (LBCPS) and a group of LAGB only (LAGBO). Groups were matched for age, gender, and pre-operative body mass index (BMI). Results LBCPS ( n = 18) had lower endpoint BMI and BMI regain percentage compared with LAGBO ( n = 54) (24.64 ± 3.76 vs. 31.0 ± 7.2 kg/m2 , p < 0.001; 13 ± 14% vs. 34 ± 31%. p < 0.001, respectively). BCPS had an independent strong protective effect for endpoint BMI regain over 25% and BMI ≥ 30 kg/m2 (HR = 0.19, p = 0.025; HR = 0.13, p = 0.046, respectively). Conclusion When compared with patients who did not undergo BCPS following LAGB, patients who underwent BCPS following LAGB had improved long-term BMI control.
The COVID-19 pandemic has led to high global demand for vaccines to safeguard public health. To that end, our institute has developed a recombinant viral vector vaccine utilizing a modified vesicular ...stomatitis virus (VSV) construct, wherein the G protein of VSV is replaced with the spike protein of SARS-CoV-2 (rVSV-ΔG-spike). Previous studies have demonstrated the production of a VSV-based vaccine in Vero cells adsorbed on Cytodex 1 microcarriers or in suspension. However, the titers were limited by both the carrier surface area and shear forces. Here, we describe the development of a bioprocess for rVSV-ΔG-spike production in serum-free Vero cells using porous Fibra-Cel
macrocarriers in fixed-bed BioBLU
320 5p bioreactors, leading to high-end titers. We identified core factors that significantly improved virus production, such as the kinetics of virus production, the use of
for oxygen supply, and medium replenishment. Implementing these parameters, among others, in a series of GMP production processes improved the titer yields by at least two orders of magnitude (2e9 PFU/mL) over previously reported values. The developed process was highly effective, repeatable, and robust, creating potent and genetically stable vaccine viruses and introducing new opportunities for application in other viral vaccine platforms.
Juvenile idiopathic arthritis (JIA) is the most common chronic rheumatic disease in children. Intra-articular corticosteroid injection (IASI), one of the cornerstones of treatment for this disease, ...is usually associated with anxiety and pain. IASI in JIA may be performed under general anesthesia, conscious sedation, or local anesthesia alone. Currently, there is no widely accepted standard of care regarding the sedation method for IASI. This review discusses the different methods of anesthesia and sedation in this procedure, emphasizing the advantages and shortcomings of each method.
The low-frequency vibrational (LFV) modes of biomolecules reflect specific intramolecular and intermolecular thermally induced fluctuations that are driven by external perturbations, such as ligand ...binding, protein interaction, electron transfer, and enzymatic activity. Large efforts have been invested over the years to develop methods to access the LFV modes due to their importance in the studies of the mechanisms and biological functions of biomolecules. Here, we present a method to measure the LFV modes of biomolecules based on Raman spectroscopy that combines volume holographic filters with a single-stage spectrometer, to obtain high signal-to-noise-ratio spectra in short acquisition times. We show that this method enables LFV mode characterization of biomolecules even in a hydrated environment. The measured spectra exhibit distinct features originating from intra- and/or intermolecular collective motion and lattice modes. The observed modes are highly sensitive to the overall structure, size, long-range order, and configuration of the molecules, as well as to their environment. Thus, the LFV Raman spectrum acts as a fingerprint of the molecular structure and conformational state of a biomolecule. The comprehensive method we present here is widely applicable, thus enabling high-throughput study of LFV modes of biomolecules.
The COVID-19 pandemic caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) increases the need for a rapid development of efficient vaccines. Among other vaccines in clinical trials, ...a recombinant VSV-∆G-spike vaccine was developed by the Israel Institute for Biological Research (IIBR) and is being evaluated. The development of an efficient downstream purification process (DSP) enables the vaccine to be advanced to clinical trials. The DSP must eliminate impurities, either process- or product-related, to yield a sufficient product with high purity, potency and quality. To acquire critical information on process restrictions and qualities, the application of in-line monitoring is vital and should significantly impact the process yield, product quality and economy of the entire process. Here, we describe an in-line monitoring technique that was applied in the DSP of the VSV-∆G-spike vaccine. The technique is based on determining the concentrations of metabolites, nutrients and a host cell protein using the automatic chemistry analyzer, Cobas Integra 400 Plus. The analysis revealed critical information on process parameters and significantly impacted purification processes. The technique is rapid, easy and efficient. Adopting this technique during the purification process improves the process yield and the product quality and enhances the economy of the entire downstream process for biotechnology and bio pharmaceutical products.
PDF
