Relationship of human cytomegalovirus-infected endothelial cells and circulating leukocytes in the pathogenesis of disseminated human cytomegalovirus infection: A narrative review.

Among the leucocyte subpopulations circulating in peripheral blood of immune-compromised patients with disseminated Human cytomegalovirus (HCMV) infection, polymorphonuclear leuckocytes (PMNL) and M/M may carry infectious virus. While only in PMNL early HCMV replicative events do occur, monocytes are susceptible to complete virus replication when they enter human organs, where as macrophages become a site of active complete virus replication. In vivo leucocytes and endothelial cells interact continuously, as suggested by several in vitro experimental findings showing the bidirectional HCMV transmission from leucocytes to and from endothelial cells with the critical aid of adhesion molecules. Recently, the neutralising antibody response in sera from subjects with primary HCMV infection was reported to be much higher and earlier than in human embryonic lung fibroblasts (HELF) cells when measured in endothelial cells and epithelial cells, where virus entry is mediated mostly by the pentamer complex gH/gL/pUL128/pUL130/pUL131, whereas it was much lower and delayed when determined in HELF, where virus entry is mediated mostly by the trimer complex gH/gL/gO. Thus, these results suggested that products of UL128L were the molecules primary responsible for the differential neutralising antibody response. This conclusion was confirmed by a series of polyclonal and monoclonal antibodies directed to the components of pUL128L. Very recently, based on two sets of experiments including inhibition and immunoblotting assays, the pentamer complex/trimer complex ratio has been finally identified as the main factor of the neutralising antibody response. This ratio may change with the virus suspension producer and target cell system as well as number of cell culture passages.

lncRNA Helf promotes hepatic inflammation and fibrosis by interacting with PTBP1 to facilitate PIK3R5 mRNA stabilization.

BACKGROUND: Hepatic fibrosis is a common consequence of chronic liver diseases without approved antifibrotic therapies. Long noncoding RNAs (lncRNAs) play an important role in various pathophysiological processes. However, the functions of certain lncRNAs involved in mediating the antifibrotic role remain largely unclear. METHODS: The RNA level of lnc-High Expressed in Liver Fibrosis (Helf) was detected in both mouse and human fibrotic livers. Furthermore, lnc-Helf-silenced mice were treated with carbon tetrachloride (CCl4) or bile duct ligation (BDL) to investigate the function of lnc-Helf in liver fibrosis. RESULTS: We found that lnc-Helf has significantly higher expression in human and mouse fibrotic livers as well as M1 polarized hepatic macrophages (HMs) and activated hepatic stellate cells (HSCs). In vivo studies showed that silencing lnc-Helf by AAV8 vector alleviates CCl4- and BDL-induced hepatic inflammation and fibrosis. Furthermore, in vitro experiments revealed that lnc-Helf promotes HSCs activation and proliferation, as well as HMs M1 polarization and proliferation in the absence or presence of cytokine stimulation. Mechanistically, our data illustrated that lnc-Helf interacts with RNA binding protein PTBP1 to promote its interaction with PIK3R5 mRNA, resulting in increased stability and activating the AKT pathway, thus promoting HSCs and HMs activation and proliferation, which augments hepatic inflammation and fibrosis. CONCLUSION: Our results unveil a lnc-Helf/PTBP1/PIK3R5/AKT feedforward, amplifying signaling that exacerbates the process of hepatic inflammation and fibrosis, thus providing a possible therapeutic strategy for hepatic fibrosis.

Hormetic effects of noncoplanar PCB exposed to human lung fibroblast cells (HELF) and possible role of oxidative stress.

Hormesis, a biphasic dose-response phenomenon, which is characterized by stimulation of an end point at a low-dose and inhibition at a high-dose. In the present study we used human lungs fibroblast (HELF) cells as a test model to evaluate the role of oxidative stress (OS) in hormetic effects of non coplanar PCB 101. Results from 3-(4,5-dime-thylthiazol-2-yl)-2,5-diphenyltetrazo-lium bromide (MTT) assay indicated that PCB101 at lower concentrations (10(-5) to 10(-1) µg mL(-1) ) stimulated HELF cell proliferation and inhibited at high concentrations (1, 5, 10, and 20 µg mL(-1) ) in a dose- and time-dependent manner. Reactive oxygen species (ROS), malondialdehyde (MDA) and superoxide dismutase (SOD) (except 48 h) showed a significant increase at higher concentrations of PCB 101 than those at the lower concentrations with the passage of time. Antioxidant enzymes such as glutathione peroxidase (GSH-Px) exhibited decreasing trends in dose and time dependent manner. Lipid peroxidation assay resulted in a significant increase (P < 0.05) of MDA level in PCB 101-treated HELF cells compared with controls, suggesting that OS plays a key role in PCB 101-induced toxicity. Comet assay indicated a significant increase in genotoxicity at higher concentrations of PCB 101 exposure compared to lower concentrations. Overall, we found that HELF cell proliferation was higher at low ROS level and vice versa, which revealed activation of cell signaling-mediated hormetic mechanisms. The results suggested that PCB 101 has hormetic effects to HELF cells and these were associated with oxidative stress.

Drought alleviation efficacy of a galactose rich polysaccharide isolated from endophytic Mucor sp. HELF2: A case study on rice plant.

Endophytes play a vital role in plant growth under biotic and abiotic stress conditions. In the present investigation, a Galactose-Rich Heteropolysaccharide (GRH) with a molecular weight of 2.98 × 105 Da was isolated from endophytic Mucor sp. HELF2, a symbiont of the East Indian screw tree Helicteres isora. OVAT (One Variable at A Time) experiment coupled with RSM (Response Surface Methodology) study exhibited 1.5-fold enhanced GRH production (20.10 g L-1) in supplemented potato dextrose broth at a pH of 7.05 after 7.5 days of fermentation in 26°C. GRH has alleviated drought stress (polyethylene glycol induced) in rice seedlings (Oryza sativa ssp. indica MTU 7093 swarna) by improving its physicochemical parameters. It has been revealed that spray with a 50-ppm dosage of GRH exhibited an improvement of 1.58, 2.38, 3, and 4 times in relative water contents and fresh weight of the tissues, root length, and shoot length of the rice seedlings, respectively "in comparison to the control". Moreover, the soluble sugars, prolines, and chlorophyll contents of the treated rice seedlings were increased upto 3.5 (0.7 ± 0.05 mg/g fresh weight), 3.89 (0.57 ± 0.03 mg/g fresh weight), and 2.32 (1,119 ± 70.8 µg/gm of fresh weight) fold respectively, whereas malondialdehyde contents decreased up to 6 times. The enzymatic antioxidant parameters like peroxidase and superoxide dismutase and catalase activity of the 50 ppm GRH treated seedlings were found to be elevated 1.8 (720 ± 53 unit/gm/min fresh weight), 1.34 (75.34 ± 4.8 unit/gm/min fresh weight), and up to 3 (100 ppm treatment for catalase - 54.78 ± 2.91 unit/gm/min fresh weight) fold, respectively. In this context, the present outcomes contribute to the development of novel strategies to ameliorate drought stress and could fortify the agro-economy of India.

Gold nanorods-mediated efficient synergistic immunotherapy for detection and inhibition of postoperative tumor recurrence.

Tumor recurrence after surgery is the main cause of treatment failure. However, the initial stage of recurrence is not easy to detect, and it is difficult to cure in the late stage. In order to improve the life quality of postoperative patients, an efficient synergistic immunotherapy was developed to achieve early diagnosis and treatment of post-surgical tumor recurrence, simultaneously. In this paper, two kinds of theranostic agents based on gold nanorods (AuNRs) platform were prepared. AuNRs and quantum dots (QDs) in one agent was used for the detection of carcinoembryonic antigen (CEA), using fluorescence resonance energy transfer (FRET) technology to indicate the occurrence of in situ recurrence, while AuNRs in the other agent was used for photothermal therapy (PTT), together with anti-PDL1 mediated immunotherapy to alleviate the process of tumor metastasis. A series of assays indicated that this synergistic immunotherapy could induce tumor cell death and the increased generation of CD3+/CD4+ T-lymphocytes and CD3+/CD8+ T-lymphocytes. Besides, more immune factors (IL-2, IL-6, and IFN-γ) produced by synergistic immunotherapy were secreted than mono-immunotherapy. This cooperative immunotherapy strategy could be utilized for diagnosis and treatment of postoperative tumor recurrence at the same time, providing a new perspective for basic and clinical research.

Activation of MAPK and Cyclin D1/CDK4 in Malignant Transformation of Human Embryonic Lung Fibroblasts Induced by Silica and Benzopyrene.

OBJECTIVE: Silica and Benzo(a)pyrene are listed as carcinogens. This study aims to explore Cyclin D1, CDK4 and difference of cell cycle adjusted by MAPK signal transduction pathway in silica and B(a)P-induced malignant transformation of human embryonic lung fibroblasts. METHODS: Activity of the subfamily (ERK, p38 and JNK) of mitogen-activated protein kinase (MAPK), cyclin D1 and CDK4 (cyclin dependent kinase) were evaluated using Human embryonic lung fibroblast (HELF) purchased from the cell room, basic research institute, Chinese Academy of Medical Sciences. The expression of cyclin D1 and CDK4 (cyclin dependent kinase) were measured in silica and B(a)P induced malignant using Western blot (WB) assay. RESULT: P-ERK and P-JNK expression increased significantly (P<0.01), while CDK4 and P-p38 expression decreased (P<0.01, P<0.05) in silica-induced malignant transformation cells compared with the control group. P-ERK, P-JNK and Cyclin D1 expression increased (P<0.01, P<0.01, P<0.05) in B(a)P-induced group compared with the control group. P-ERK and P-JNK expression decreased (P<0.01), while P-p38, Cyclin D1 and CDK4 expression increased (P<0.05, P<0.05, P<0.01) in B(a)P-induced group compared with the silica-induced group. CONCLUSION: MAPK and cyclin D1/CDK4 activation expressed differently in human embryo lung fibroblasts malignant transformation induced by silica and benzopyrene.

Anti-fibrotic effect of melittin on TRIM47 expression in human embryonic lung fibroblast through regulating TRIM47 pathway.

AIMS: To investigate the effect and underlying mechanism of melittin and tripartite motif (TRIM) family in human embryonic lung fibroblast (HELF). MATERIALS AND METHODS: Lentiviral RNA interference vector and lentiviral overexpression vector were constructed and packaged by transfecting 293T cells; the proliferation of HELF was examined using Cell Counting Kit 8; Western blot and qRT-PCR were performed to examine protein and mRNA expression; the interaction with protein phosphatase magnesium-dependent 1A (PPM1A) was examined by Co-immunoprecipitation. KEY FINDINGS: Compared with the control group, the mRNA expression of the TRIM6, TRIM8 and TRIM47 in the IPF group significantly increased. Melittin inhibited the mRNA expression and protein expression levels of TRIM47, the HELF proliferation, the hydroxyproline levels, and the phosphorylation of Smad2/3; the interference of TRIM47 inhibited the protein expression of Vimentin, α-SMA, CTGF, the phosphorylation of Smad2/3 and the synthesis of hydroxyproline; TRIM47 overexpression elevated the phosphorylation of Smad2/3, induced ubiquitination of PPM1A and decreased the expression level of PPM1A, while TRIM47 RNA interference reversed this result. SIGNIFICANCE: Melittin has anti-fibrotic effect in HELF by directly reducing the phosphorylation of Smad2/3 or indirectly reducing the phosphorylation of Smad2/3 by decreasing the expression levels of TRIM47 whose overexpression induces ubiquitination of PPM1A.

Thalidomide inhibits the gene promoter of connective tissue growth factor in human embryonic lung fibroblasts.

BACKGROUND: The etiology and pathogenesis of idiopathic pulmonary fibrosis (IPF) remain unclear, and the early detection and treatment are vital to the prognosis of IPF patients. It's necessary to investigate the effect of thalidomide on the gene promoter activation of connective tissue growth factor (CTGF) induced by transforming growth factor-ß1 (TGF-ß1) in human embryonic lung fibroblast (HELF). METHODS: The gene vector of pGL3-CTGFP containing human CTGF gene promoter was constructed and transfected into HELF cells. We used different TGF-ß1 (0, 2.5, 5, 10 and 20 µg/L respectively) to stimulate HELF to identify the optimal concentration for gene promoter of CTGF. The activity of luciferase was measured to observe the effect of TGF-ß1 and THALIDOMIDE on the activity of CTGF gene promoter. RESULTS: The relative luciferase activity increased significantly with the stimulation of TGF-ß1, and the relative luciferase activity peaked in the 5 µg/L TGF-ß1 group (all P<0.01). The thalidomide inhibited the TGF-ß1-induced activation of CTGF gene promoter in HELF, and the effect peaked in the 25 µg/L group (all P<0.001). CONCLUSIONS: Thalidomide produces a significant inhibitory effect on the gene promoter activation of CTGF induced by TGF-ß1 in HELF, it may be a potentially effective drug for the treatment of pulmonary fibrosis.

PCB118-Induced Cell Proliferation Mediated by Oxidative Stress and MAPK Signaling Pathway in HELF Cells.

The present study used human lung fibroblast (HELF) cells as a test model to evaluate the role of oxidative stress (OS) and extracellular signal-regulated kinases 1/2 (ERK1/2) protein in HELF cell proliferation exposed to PCB118. Results from 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide demonstrated that PCB118 at lower concentrations stimulated proliferation of HELF cell and abrogate proliferative effect at higher dose concentrations and in a time-dependent manner. Moreover, reactive oxygen species, malondialdehyde (MDA), and superoxide dismutase showed a significant increase at higher concentrations of PCB118 than the lower concentrations with the passage of time. Antioxidant enzymes such as glutathione peroxidase exhibited decreasing trends in dose- and time-dependent manner. Lipid peroxidation assay resulted in a significant increase in MDA level in PCB118-treated HELF cells compared with controls, suggesting that OS plays a key role in PCB118-induced toxicity. Comet assay indicated a significant increase in genotoxicity at higher concentrations of PCB118 exposure than the lower concentrations. It was found that PCB118 showed expression of ERK1/2 protein after 4 hours, while after 48 hours, the protein expression was less, indicating PCB toxicity to MAPK protein of HELF cell. Oxidative stress, ERK1/2, and HELF cell proliferation exhibited correlation. The results will elaborate toxicological evaluation of PCB118 to HELF cells and will help to develop drug for PCB-induced diseases.

Os marcos legais das políticas de medicamentos no Brasil contemporâneo (1990-2006) / The legal landmarks of the drugs policies in contemporary Brazil (1990-2006)

O presente trabalho tem como objeto o processo de reorientação da assistência farmacêutica levada a termo no Brasil a partir da Lei 8.080 de 1990. Essa lei é o primeiro marco legal de um processo que gerou distintas legislações relativas à assistência farmacêutica no âmbito do SUS. O objetivo principal do trabalho foi levantar, sistematizar e nalisar os principais documentos e propostas que promoveram a reorientação da assistência farmacêutica desde 1990 até o presente. O foco do trabalho foi a elaboração, em 1998, da Política Nacional de Medicamentos (PNM), seus antecedentes Central de Medicamentos – (CEME) – Decreto 68.806/71 e a Lei 6.360/76 relativa às ações de vigilância sanitária, bem como a crescente edição de legislações voltadas para a sua efetiva implantação: a criação da Agência Nacional de Vigilância Sanitária – (ANVISA) (1999); a Lei 9.787, conhecida como a Lei dos Genéricos (1999), e a Política Nacional de Assistência Farmacêutica (2004). O trabalho apresenta uma periodização da trajetória das políticas de medicamentos no Brasil contemporâneo e procura assinalar algumas das características desse processo. Entre essas características figura o caráter abrangente dessas legislações, bem como a insuficiente articulação entre estas no sentido de promover a consolidação de uma nova cultura de medicamentos no país. Este trabalho não se limitou a construir uma visão retrospectiva do processo de reorientação da assistência farmacêutica em curso no Brasil contemporâneo, mas procurou buscar alguns elementos para a formação de uma visão prospectiva - na qual estudos avaliativos ganham importância estratégica – das ações no campo da assistência farmacêutica.

The present work focuses on the process of the reorientation of pharmaceutical assistance introduced in Brazil with Law 8,080 of 1990. This law is the first legal landmark of a process that created distinct legal provisions concerning pharmaceutical assistance in the setting of SUS (the Unified National Health System). The main purpose of the study was identifying, systematizing and analyzing the principal documents and proposals that promoted the reorientation of pharmaceutical assistance from 1990 to today. The focus of the work was the elaboration, in 1998, of the National Drug Policy, its antecedents Central for Drugs - (CEME) - Decree 68,806/71 and Law 6,360/76 relative to public health surveillance activities, as well as the increasing number of legislative provisions focused on its effective implementation: establishment: the creation of the National Health Surveillance Agency - (ANVISA) (1999); Law 9,787, known as the Generic Drug Law (1999) and the National Pharmaceutical Assistance Policy (2004). The study presents a timeline of the trajectory of drug policy in Brazil today and seeks to highlight some of the characteristics of this process. Among these characteristics there is the all-inclusive nature of these pieces of legislation as well as the insufficient coordination between them that would promote the consolidation of a new medication culture in the country. This work was not limited to a retrospective view of the reorientation process of pharmaceutical assistance in practice today in Brazil, but it tried to find some elements for the formation of a prospective vision, in which evaluation studies become an important strategy in the policies of pharmaceutical assistance.