Some observations on the effect of Daflon (micronized purified flavonoid fraction of Rutaceae aurantiae) in bancroftian filarial lymphoedema

BACKGROUND: Morbidity management is a core component of the global programme for the elimination of lymphatic filariasis. In a double-blind clinical trial, the tolerability and efficacy of Daflon (500 mg) + DEC (25 mg) or DEC (25 mg) alone, twice daily for 90 days, was studied in 26 patients with bancroftian filarial lymphoedema. RESULTS: None of the patients in either drug group reported any adverse reaction throughout the treatment period (90 days). Haematological and biochemical parameters were within normal limits and there was no significant difference between the pre-treatment (day 0) and post-treatment (day 90) values. The group receiving Daflon showed significant reduction in oedema volume from day 90 (140.6 PlusMinus; 18.8 ml) to day 360 (71.8 PlusMinus; 20.7 ml) compared to the pre-treatment (day 0, 198.4 PlusMinus; 16.5 ml) value. This accounted for a 63.8% reduction in oedema volume by day 360 (considering the pre-treatment (day 0) as 100%). In the DEC group, the changes in oedema volume (between day 1 and day 360) were not significant when compared to the pre-treatment (day 0) value. The percentage reduction at day 360 was only 9%, which was not significant (P > 0.05). CONCLUSION: This study has shown that Daflon (500 mg, twice a day for 90 days) is both safe and efficacious in reducing oedema volume in bancroftian filarial lymphoedema. Further clinical trials are essential for strengthening the evidence base on the role of this drug in the morbidity management of lymphatic filariasis.     

Direct intracardiac injection of umbilical cord-derived stromal cells and umbilical cord blood-derived endothelial cells for the treatment of ischemic cardiomyopathy

The development of new therapeutic strategies is necessary to reduce the worldwide social and economic impact of cardiovascular disease, which produces high rates of morbidity and mortality. A therapeutic option that has emerged in the last decade is cell therapy. The aim of this study was to compare the effect of transplanting human umbilical cord-derived stromal cells (UCSCs), human umbilical cord blood-derived endothelial cells (UCBECs) or a combination of these two cell types for the treatment of ischemic cardiomyopathy (IC) in a Wistar rat model. IC was induced by left coronary artery ligation, and baseline echocardiography was performed seven days later. Animals with a left ventricular ejection fraction (LVEF) of ≤40% were selected for the study. On the ninth day after IC was induced, the animals were randomized into the following experimental groups: UCSCs, UCBECs, UCSCs plus UCBECs, or vehicle (control). Thirty days after treatment, an echocardiographic analysis was performed, followed by euthanasia. The animals in all of the cell therapy groups, regardless of the cell type transplanted, had less collagen deposition in their heart tissue and demonstrated a significant improvement in myocardial function after IC. Furthermore, there was a trend of increasing numbers of blood vessels in the infarcted area. The median value of LVEF increased by 7.19% to 11.77%, whereas the control group decreased by 0.24%. These results suggest that UCSCs and UCBECs are promising cells for cellular cardiomyoplasty and can be an effective therapy for improving cardiac function following IC.     

Airway hyperresponsiveness is associated with airway remodeling but not inflammation in aging Cav1-/- mice

Background

Airway inflammation and airway remodeling are the key contributors to airway hyperresponsiveness (AHR), a characteristic feature of asthma. Both processes are regulated by Transforming Growth Factor (TGF)-β. Caveolin 1 (Cav1) is a membrane bound protein that binds to a variety of receptor and signaling proteins, including the TGF-β receptors. We hypothesized that caveolin-1 deficiency promotes structural alterations of the airways that develop with age will predispose to an increased response to allergen challenge.

Methods

AHR was measured in Cav1-deficient and wild-type (WT) mice 1 to 12 months of age to examine the role of Cav1 in AHR and the relative contribution of inflammation and airway remodeling. AHR was then measured in Cav1^-/- and WT mice after an ovalbumin-allergen challenge performed at either 2 months of age, when remodeling in Cav1^-/- and WT mice was equivalent, and at 6 months of age, when the Cav1^-/- mice had established airway remodeling.

Results

Cav1^-/- mice developed increased thickness of the subepithelial layer and a correspondingly increased AHR as they aged. In addition, allergen-challenged Cav1^-/- mice had an increase in AHR greater than WT mice that was largely independent of inflammation. Cav1^-/- mice challenged at 6 months of age have decreased AHR compared to those challenged at 2 months with correspondingly decreased BAL IL-4 and IL-5 levels, inflammatory cell counts and percentage of eosinophils. In addition, in response to OVA challenge, the number of goblet cells and α-SMA positive cells in the airways were reduced with age in response to OVA challenge in contrast to an increased collagen deposition further enhanced in absence of Cav1.

Conclusion

A lack of Cav1 contributed to the thickness of the subepithelial layer in mice as they aged resulting in an increase in AHR independent of inflammation, demonstrating the important contribution of airway structural changes to AHR. In addition, age in the Cav1^-/- mice is a contributing factor to airway remodeling in the response to allergen challenge.     

Regulatable gutless adenovirus vectors sustain inducible transgene expression in the brain in the presence of an immune response against adenoviruses

In view of recent serious adverse events and advances in gene therapy technologies, the use of regulatable expression systems is becoming recognized as indispensable adjuncts to successful clinical gene therapy. In the present work we optimized high-capacity adenoviral (HC-Ad) vectors encoding the novel tetracycline-dependent (TetOn)-regulatory elements for efficient and regulatable gene expression in the rat brain in vivo. We constructed two HC-Ad vectors encoding beta-galactosidase (beta-gal) driven by a TetOn system containing the rtTAS(s)M2 transactivator and the tTS(Kid) repressor under the control of the murine cytomegalovirus (mCMV) (HC-Ad-mTetON-beta-Gal) or the human CMV (hCMV) promoter (HC-Ad-hTetON-beta-Gal). Expression was tightly regulatable by doxycycline (Dox), reaching maximum expression in vivo at 6 days and returning to basal levels at 10 days following the addition or removal of Dox, respectively. Both vectors achieved higher transgene expression levels compared to the expression from vectors encoding the constitutive mCMV or hCMV promoter. HC-Ad-mTetON-beta-Gal yielded the highest transgene expression levels and expressed in both neurons and astrocytes. Antivector immune responses continue to limit the clinical use of vectors. We thus tested the inducibility and longevity of HC-Ad-mediated transgene expression in the brain of rats immunized against adenovirus by prior intradermal injections of RAds. Regulated transgene expression from HC-Ad-mTetON-beta-Gal remained active even in the presence of a significant systemic immune response. Therefore, these vectors display two coveted characteristics of clinically useful vectors, namely their regulation and effectiveness even in the presence of prior immunization against adenovirus.     

Novel fiber-dependent entry mechanism for adenovirus serotype 5 in lacrimal acini

The established mechanism for infection of most cells with adenovirus serotype 5 (Ad5) involves fiber capsid protein binding to coxsackievirus-adenovirus receptor (CAR) at the cell surface, followed by penton base capsid protein binding to alpha(v) integrins, which triggers clathrin-mediated endocytosis of the virus. Here we determined the identity of the capsid proteins responsible for mediating Ad5 entry into the acinar epithelial cells of the lacrimal gland. Ad5 transduction of primary rabbit lacrimal acinar cells was inhibited by excess Ad5 fiber or knob (terminal region of the fiber) but not excess penton base. Investigation of the interactions of recombinant Ad5 penton base, fiber, and knob with lacrimal acini revealed that the penton base capsid protein remained surface associated, while the knob domain of the fiber capsid protein was rapidly internalized. Introduction of rabbit CAR-specific small interfering RNA (siRNA) into lacrimal acini under conditions that reduced intracellular CAR mRNA significantly inhibited Ad5 transduction, in contrast to a control (nonspecific) siRNA. Preincubation of Ad5 with excess heparin or pretreatment of acini with a heparinase cocktail each inhibited Ad5 transduction by a separate and apparently additive mechanism. Functional and imaging studies revealed that Ad5, fiber, and knob, but not penton base, stimulated macropinocytosis in acini and that inhibition of macropinocytosis significantly reduced Ad5 transduction of acini. However, inhibition of macropinocytosis did not reduce Ad5 uptake. We propose that internalization of Ad5 into lacrimal acini is through a novel fiber-dependent mechanism that includes CAR and heparan sulfate glycosaminoglycans and that the subsequent intracellular trafficking of Ad5 is enhanced by fiber-induced macropinocytosis.     

A Rat Brain cDNA Encodes Enzymatically Active GABA Transaminase and Provides a Molecular Probe for GABA-Catabolizing Cells

Abstract: cDNAs encoding γ-aminobutyric acid aminotransferase (GABA-T) were isolated from a λZAP rat hippocampal cDNA expression library by two independent cloning methods, immunological screening with an antimouse GABA-T antibody and plaque hybridization with a GABA-T cDNA probe derived by polymerase chain reaction. We have produced enzymatically active GABA-T from a rat brain cDNA containing the full-length GABA-T coding region. Our rat brain GABA-T cDNAs hybridize to mRNAs in brain and peripheral tissues, including liver, kidney, and testis. We have also detected GABA-T mRNA in GABAergic cells of rat cerebellar cortex by in situ hybridization. Our rat brain GABA-T probe hybridizes to Purkinje, basket, stellate, and Golgi II cells, the same GABAergic neurons previously shown to contain glutamate decarboxylase GAD₈₅ and GAD₈₇.     

Tolerability and efficacy of single dose albendazole,diethylcarbamazine citrate (DEC) or co-administration of albendazole with DEC in the clearance of <Emphasis Type="Italic">Wuchereria bancrofti</Emphasis>in asymptomatic microfilaraemic volunteers in Pondicherry,South India: a hospital-based study

Background  

The tolerability and efficacy of single dose albendazole (400 mg), diethylcarbamazine citrate (DEC) (6 mg/kg bodyweight) or co-administration of albendazole (400 mg) + DEC (6 mg/kg bodyweight) was studied in 54 asymptomatic Wuchereria bancrofti microfilaraemic volunteers in a double blind hospital-based clinical study.     

Assessing the binding and endocytosis activity of cellular receptors using GFP-ligand fusions

We have developed a simple scheme for characterizing ligand-receptor binding and post-binding activity on living cells. Our approach makes use of green fluorescent protein (GFP) as an auto-fluorescent tag to label protein ligands. We have constructed GFP-tagged ligands that can be expressed in bacteria as soluble fusion proteins. A cell-binding assay using fluorescence-activated cell sorting (FACS) demonstrates that GFP-tagged proteins retain their wild-type receptor-binding specificity. Using this assay, we measure ligand binding on unfixed cells and demonstrate receptor specificity using specific competitors. To determine the ability of receptor targets to internalize, we developed a second FACS-based assay to detect the rate and percentage of internalized ligand in living cells. Noninternalizing control ligands and fluorescence microscopy of treated cells confirm that our assay is reliable for determining receptor internalization activity.     

Antagonist-mediated down-regulation of toll-like receptors increases the prevalence of human papillomavirus infection in systemic lupus erythematosus

IntroductionPrevalence of an abnormal Papanicolaou smear was significantly increased in lupus patients in cross-sectional studies, associated with a higher prevalence of high-risk human papillomavirus (HPV) infection. The nucleic acid-specific Toll-like receptors (TLRs) locate at the endolysosomal compartments and trigger the induction of cytokines for the innate immune response. This study evaluated whether abnormal host innate immune response in lupus patients may enhance HPV persistence.MethodsProtein levels of TLRs 3, 7, 8 and 9 in cervical epithelial cells of lupus patients and controls with or without HPV infection were assessed using flow cytometry. Characteristics associated with the differential expression of TLRs in systemic lupus erythematosus (SLE) were elucidated. The effect and interferon-stimulated genes (ISGs) (ISG15 and Mx-1) gene expressions were then measured in oncogenic HeLa (HPV18), CaSki (HPV) and C33A (HPV negative) cell lines using flow cytometry and quantitative real-time PCR. Ex vivo productions of cytokines and interferon-gamma (IFN-γ) upon TLR ligands stimulations were subsequently measured using cytometric bead array and ELISA.ResultsFor subjects with HPV infection, levels of TLR3 and TLR7 were significantly lower in lupus patients compared with controls. Significantly decreased TLRs 7, 8 and 9 levels were observed in HPV-negative SLE compared to healthy controls. For SLE with and without HPV infection, TLR7 and 9 levels were significantly lower in infected SLE than those in HPV-negative patients. Independent explanatory variables associated with down-regulation of TLR7 level included HPV infection and a higher cumulative dose of prednisolone; while a higher cumulative dose of hydroxychloroquine and HPV infection were associated with down-regulation of TLR9 level. In cervical cell lines, TLRs 3, 7, 8, 9 protein levels and antiviral ISG15 and Mx-1 gene expressions were inhibited in two oncogenic HPV types. Functional data showed that the induction of pro-inflammatory cytokines by TLR ligands (R837, ssRNA and ODN2395) was greatly impaired in CaSki and HeLa than C33A cells.ConclusionsIn conclusion, prednisolone and TLR antagonist (hydroxychloroquine) may down-regulate protein levels of TLR7 and TLR9 in lupus patients, thereby decreasing the innate immune response against HPV infection. Upon infection, HPV further down-regulate TLR7 and 9 levels for viral persistence. Furthermore, reduction of nucleic acid-sensing TLRs 7, 8 and 9 in carcinogenic HPVs ensures that the expression of inducible pro-inflammatory cytokines is minimized to prevent the expression of antiviral ISGs (ISG15 and Mx-1) on a biologically relevant antiviral response.