Antitumor and immunomodulatory activity of fucoidan from the brown alga Lessonia trabeculata (Lessoniaceae) on breast cancer spheroids / Actividad antitumoral e inmunomoduladora de fucoidan del alga parda Lessonia trabeculata (Lessoniaceae) en esferoides de cáncer de mama

Abstract Introduction: The therapeutic benefits of the brown algae fucoidan in the treatment of breast cancer have attracted considerable interest in recent years. However, research using spheroids which provide relevant results in trials for antitumor and immunomodulatory products because they adequately simulate the tumor microenvironment, is limited. Objective: To evaluate the antitumor and immunomodulatory activity of Lessonia trabeculata fucoidan (LtF), native to the Peruvian Sea, on two types of multicellular tumor spheroids. Methods: The study was conducted from January to December 2021. Two types of spheroides were elaborated: from 4T1 tumor cells (MTS), and from 4T1 tumor cells+mouse splenocytes (MTSs). The antitumor activity of LtF was evaluated in MTS by quantifying cell viability with MTT. Immunomodulatory activity was determined in MTSs using the IC50 for two types of treatment: simple, fucoidan alone (LtF) and combined, fucoidan+doxorubicin (LtF+Dox). Pro-inflammatory (TNF-α, IL-6) and anti-inflammatory (IL-10, TGF-β) cytokine production was quantified by sandwich ELISA 72 h after treatment. Dox was used as positive control in all assays. Results: LtF exerted antitumor activity as evidenced by increased necrotic zone and cell debris formation compared to the untreated control. Antitumor activity was concentration dependent between 100 and 6 000 μg/ml. In MTSs, simple treatment increased IL-6 and decreased IL-10 and TGF-β production. The combined treatment significantly reduced TGF-β production. In both treatments and Dox, there was an increase in IL-6 compared to the untreated control. The highest production of IL-10 and TGF-β was observed in the untreated control, compatible with a highly immunosuppressive tumor microenvironment. Conclusions: LtF is a good candidate for the treatment of breast cancer and can immunomodulate the tumor microenvironment alone or in combination with Dox.

Resumen Introduccción: Los beneficios terapéuticos del fucoidan de algas pardas en el tratamiento del cáncer de mama han despertado gran interés en los últimos años. Sin embargo, las investigaciones con esferoides son limitadas, éstos proporcionan resultados relevantes en ensayos de productos antitumorales e inmunomoduladores porque simulan adecuadamente el microambiente tumoral. Objetivo: Evaluar la actividad antitumoral e inmunomoduladora del fucoidan de Lessonia trabeculata (LtF), nativa del Mar Peruano, en dos tipos de esferoides tumorales multicelulares. Métodos: El estudio se realizó de enero a diciembre de 2021. Se elaboraron dos tipos de esferoides: con células tumorales 4T1 (MTS) y con células tumorales 4T1+esplenocitos de ratón (MTSs). La actividad antitumoral de LtF se evaluó en MTS cuantificando la viabilidad celular con MTT. La inmunomodulación se determinó en MTSs utilizando la IC50 para dos tipos de tratamiento: simple, fucoidan solo (LtF) y combinado, fucoidan+doxorubicina (LtF+Dox). La producción de citoquinas proinflamatorias (TNF-α, IL-6) y antiinflamatorias (IL-10, TGF-β) se cuantificó mediante ELISA sándwich 72 h post-tratamiento. En todos los ensayos se utilizó Dox como control positivo. Resultados: En los MTS, el LtF ejerció actividad antitumoral evidenciada por aumento de la zona necrótica y formación de restos celulares respecto al control no tratado. La actividad antitumoral fue concentración-dependiente entre 100 y 6 000 μg/ml. En los MTSs, con el tratamiento simple se incrementó IL-6 y disminuyeron IL-10 y TGF-β. El tratamiento combinado redujo significativamente la producción de TGF-β. Los dos tratamientos y Dox incrementaron IL-6 respecto al control no tratado. La mayor producción de IL-10 y TGF-β se observó en los no tratados, compatible con un microambiente tumoral altamente inmunosupresor. Conclusiones: El LtF es un buen candidato para tratar el cáncer de mama y puede inmunomodular el microambiente tumoral solo o en combinación con Dox.

Mecanismos y dianas terapéuticas del inflamasoma NLRP3 en la miocardiopatía diabética / NLRP3 inflammasome mechanism and therapeutic targets in diabetic cardiomyopathy

Resumen La cardiomiopatía diabética es una complicación grave de la diabetes causada por estrés oxidativo, inflamación, resistencia a la insulina, fibrosis miocárdica y lipotoxicidad. Se trata de un padecimiento insidioso, complejo y difícil de tratar. El inflamasoma NLRP3 desencadena la maduración y liberación de citoquinas proinflamatorias, participa en procesos fisiopatológicos como la resistencia a la insulina y la fibrosis miocárdica, además de estar estrechamente relacionado con la aparición y progresión de la cardiomiopatía diabética. El desarrollo de inhibidores dirigidos a aspectos específicos de la inflamación sugiere que el inflamasoma NLRP3 puede utilizarse para tratar la cardiomiopatía diabética. Este artículo pretende resumir el mecanismo y las dianas terapéuticas del inflamasoma NLRP3 en la cardiomiopatía diabética, así como aportar nuevas sugerencias para el tratamiento de esta enfermedad.

Abstract Diabetic cardiomyopathy (DCM) is a serious complication of diabetes caused by oxidative stress, inflammation, insulin resistance, myocardial fibrosis, and lipotoxicity; its nature is insidious, complex and difficult to treat. NLRP3 inflammasome triggers the maturation and release of pro-inflammatory cytokines, participates in pathophysiological processes such as insulin resistance and myocardial fibrosis, in addition to being closely related to the development and progression of diabetic cardiomyopathy. The development of inhibitors targeting specific aspects of inflammation suggests that NLRP3 inflammasome can be used to treat diabetic cardiomyopathy. This paper aims to summarize NLRP3 inflammasome mechanism and therapeutic targets in diabetic cardiomyopathy, and to provide new suggestions for the treatment of this disease.

Anti-inflammatory and antioxidant potential of Artemisia dracunculus L. aqueousextract against acetic acid induced ulcerative colitis in male Wistar rats

Ulcerative colitis (UC) is one of the two types of inflammatory bowel disease (IBD) which is increasing worldover due to modern life style. Patients with UC are prone to develop colorectal cancer. While the disease severity decides the treatment option, researchers look towards herbal medicines with anti-inflammatory properties for minimal or nil side effects. Artemisia dracunculus L., commonly called Tarragon, is a medicinal herb used in traditional Asian medicine mainly in Iran, India, Pakistan and Azerbaijan due to its special compounds. In this study, we tried to elucidate the effects of aqueous extract of tarragon on acetic acid induced ulcerative colitis (UC) in rats. Male Wistar rats were grouped into four groups of eight each viz., control; experimental control (UC was induced via luminal instillation of 4% acetic acid); and UC induced + aqueous tarragon extract (100 mg/kg) or prednisolone (2 mg/kg) orally for ten consecutive days. Tissue specimens were collected after the experimental period for evaluation of caspase-3 and cyclooxygenase-2 (COX-2) expression by immunohistochemistry. Real-time PCR was used to monitor the levels of IL-1, IL-6 and TNF-? in colonic homogenates. Moreover, the levels of myeloperoxidase, nitric oxide and total antioxidant capacity were measured in colonic homogenates. The results showed that both treatment regimens could similarly reduce the severity of disease symptoms. Treatment with aqueous extract of tarragon caused a better improvement (P <0.05) in the levels of myeloperoxidase enzyme, and total antioxidant capacity of colonic homogenates compared to prednisolone. Nevertheless, the levels of the expression of caspase-3, and COX-2 and TNF-? were reduced in UC rats received prednisolone more than UC rats received aqueous extract of tarragon. The was no statistical difference in the levels of nitric oxide, IL-1 and IL-6 between UC rats received tarragon extract or prednisolone. Overall, these findings suggest that the aqueous extract of tarragon is a promising strategy to control ulcerative colitis. Aqueous extract can also be used as an anti-inflammatory and immune system stimulant in conditions where the immune system is damaged.

Chlorogenic acid derived from Moringa oleifera leaf as a potential antiinflammatory agent against cryptosporidiosis in mice

@#Cryptosporidiosis is a serious illness in immunodeficient patients, and there is still no drug that can completely remove the parasite from the host. The present study represents the first report investigating the impact of the active molecule chlorogenic acid (CGA), naturally isolated from Moringa oleifera leaf extract (EMOLE), on immunosuppressed, Cryptosporidium parvum-infected BALB/c mice. Mice were divided into five groups: normal mice, infected immunosuppressed mice, and infected immunosuppressed mice treated with EMOLE, CGA, and nitazoxanide (NTZ) drugs. Parasitological, immunological, and histopathological investigations were recorded besides differences in the mice’ body weight. Infected control mice showed elevated levels of oocyst shedding throughout the study. The EMOLE- and CGA-treated groups showed 84.2% and 91.0% reductions in oocyst shedding, respectively, with no significant difference compared to the drug control. The inflammatory markers IFN-γ, IL-6, IL-1β, and TNF-α were significantly higher in the infected control group. Treatment with 300 mg/kg/day of EMOLE or 30 mg/kg/day of CGA significantly downregulated pro-inflammatory cytokine levels compared to the infected group, although they did not change significantly compared to the NTZ-treated group. Histopathology of intestinal sections showed inflammatory and pathological changes in the infected control group. Low-grade tissue changes and an obvious improvement in villi structure were seen in mice treated with CGA. This study highlighted the role of CGA, isolated and purified from EMOLE, as an effective anti-inflammatory agent in eradicating C. parvum infection.

Astaxanthin protects against hearing impairment in diabetic rats

Abstract Objectives: Diabetes Mellitus (DM) causes an increase in oxidative stress that leads to deterioration in auditory functions. Astaxanthine (AST) is known to have strong antioxidant effects. In this study, the aim is to investigate the effect of AST against hearing loss that is due to DM. Methods: This study is an experimental animal study. The study was designed in four groups with 8 animals (n = 8) in each group. The groups were as follows; Control Group (CNT), Diabetic Group (DM), AST applied diabetic group (DM+AST), and AST applied non-diabetic group (AST). Streptozotocin was applied in rats to induce DM. AST was administered by oral gavage. Auditory Brainstem Responses (ABR) and Distortion Product Otoacoustic Emissions (DPOAE) tests were performed on several days of the study. At the end of the study, pro-inflammatory cytokine levels were analyzed in cochlear tissue samples, and Glutathione Peroxidase (GPx), Superoxide Dismutase (SOD), Catalase (CAT) and Malondialdehyde (MDA) levels were measured. Results: When the findings obtained in the ABR and DPOAE tests in the DM group, it was observed that there was a significant deterioration in the hearing sense. This deterioration was not observed in the DM+AST group. In the DM group, GPx, SOD and CAT levels decreased and MDA levels increased in blood and cochlear tissue. Compared to the DM group, it was noted that antioxidant enzyme levels increased and MDA levels decreased in the DM+AST group. Cochlear tissue pro-inflammatory cytokine levels, which increased with DM, were significantly decreased in the DM+AST group. Conclusion: Even though the effects of AST were investigated in a diabetic experimental animal model, if this molecule is proven to be effective in diabetic humans, it can be considered an adjunct therapeutic option with its antioxidant effects. Level of evidence: The level of evidence of this article is 5. This article is an experimental animal and laboratory study.

Efecto del extracto hidroalcohólico de la planta Lampaya medicinalis Phil . sobre marcadores inflamación inducidos por ácidos grasos en hepatocitos humanos HepG2

RESUMEN Introducción: la enfermedad de hígado graso no alcohólico (EHGNA) se caracteriza por la acumulación de gotas lipídicas (GL) y sobre expresión de la proteína de GL Perilipina 1 (PLIN1) en los hepatocitos. En su patogénesis y progresión participan NF-ĸB, caspasa-1 y citoquinas proinflamatorias como IL-1β. La medicina popular del norte de Chile utiliza la planta Lampaya medicinalis Phil. (Verbenaceae) contra enfermedades. Objetivo: evaluar el efecto de un extracto hidroalcohólico de lampaya (EHL) sobre la expresión de marcadores inflamatorios y proteínas asociadas a las GL en hepatocitos tratados con ácidos grasos. Métodos: se incubó hepatocitos HepG2 con 0,66 mM de ácido oleico (AO) y 0,33 mM de ácido palmítico (AP) por 24 o 48 horas en presencia o no de EHL. Se evaluó la expresión proteica de NF-ĸB, PLIN1 y caspasa-1 por Western blot y la expresión de ARNm de IL-1β por qPCR. Resultados: los hepatocitos tratados por 48 h con AO/AP mostraron un aumento en la expresión de IL-1β que fue revertido por la co-incubación con EHL. Conclusión: estos antecedentes aportan nueva evidencia respecto a la actividad biológica del EHL en un modelo de alteraciones metabólicas e inflamatorias, asociadas a la EHGNA, inducidas por AO/AP en hepatocitos humanos.

ABSTRACT Introduction: Nonalcoholic fatty liver disease (NAFLD) is characterized by the accumulation of lipid droplets (LD) and overexpression of the LD-associated protein Perilipin 1 (PLIN1). NF-ĸB, caspase-1 and proinflammatory cytokine such as IL-1β participate in the pathogenesis and progression of NAFLD. Traditional medicine in northern Chile uses the plant Lampaya medicinalis Phil. (Verbenaceae) against diseases. Objective: To evaluate the effect of a hydroalcoholic extract of lampaya (HEL) on the expression of inflammatory markers and LD-associated proteins in hepatocytes treated with fatty acids. Methods: HepG2 hepatocytes were incubated with 0.66 mM oleic acid (OA) and 0.33 mM palmitic acid (PA) for 24 or 48 h in the presence or not of HEL. The protein expression of NF-ĸB, PLIN1 and caspase-1 was evaluated by Western blot while the mRNA expression of IL-1β was assessed by qPCR. Results: hepatocytes treated for 48 h with OA/AP showed an increase in IL-1β expression that was reversed by co-incubation with HEL. Conclusion: These antecedents provide new evidence regarding the biological activity of HEL in a model of metabolic and inflammatory alterations, associated with NAFLD, induced by OA/PA in human hepatocytes.

Polissacarídeos de Lentinula edodes suprimem a expressão de citocinas pró-inflamatórias e de colite em camundongos / Lentinula edodes polysaccharides suppressed pro-inflammatory cytokines expression and colitis in mice

ABSTRACT Background: Polysaccharides from edible mushrooms possess immunomodulatory, anti-inflammatory, and anti-tumor activities. Recent studies indicated that necroptosis plays a role in the pathogenesis of inflammatory diseases and mediates increased expression of inflammatory cytokines. Objective: Therefore, it is imperative to determine the impact of polysaccharide extract from Lentinula edodes (L. edodes) on inflammatory cytokines in experimental model of colitis in mice. Methods: Female C57BL/6 mice divided into three or four mice per group were used for this study. Polysaccharide sample was orally administered to mice prior to (7 days) and during colitis induction with 2.5% dextran sodium sulfate (7 days), followed by additional 3 days of administration. Changes in body weight and colon length were used as markers for colitis, and pro-inflammatory cytokines and tumor necrosis factor receptor 1 (TNFR1) expressions, as well as necroptosis were analyzed in the colon of colitis mice. Data obtained were analysed by Tukey-Kramer and two-tailed standard t tests. Results: The results indicated that the polysaccharide sample suppressed colitis in mice using effects on the body weight and colon length as markers. Also, it was demonstrated that necrostatin-1, a specific inhibitor of necroptosis, suppressed the expression of interleukin (IL)-8, a pro-inflammatory chemokine, in Caco-2 cells induced necroptosis induced by zVAD and TNF-α, an indication that necroptosis may be involved in the expression of pro-inflammatory cytokines. Moreover, the polysaccharide sample suppressed the expression of pro-inflammatory cytokines such as tumor necrosis factor (TNF)-α, IL-6, IL-1β, and interferon (IFN)-γ in the colon of mice. Conclusion: These results suggested that the suppressive effects of the polysaccharide sample on inflammatory cytokines expression may contribute to its anti-colitis effect, and so may serve as a potent therapeutic agent against inflammatory bowel disease.

RESUMO Contexto: Polissacarídeos de cogumelos comestíveis possuem atividades imunomodulatórias, anti-inflamatórias e anti-tumorais. Estudos recentes indicaram que a necroptose desempenha um papel na patogênese de doenças inflamatórias e regula o aumento da expressão de citocinas inflamatórias. Objetivo: Torna-se imprescindível determinar o impacto do extrato de polissacarídeo de Lentinula edodes (L. edodes) em citocinas inflamatórias em modelo experimental de colite em camundongos. Métodos: Foram utilizados para este estudo os camundongos C57BL/6 femininos divididos em três ou quatro camundongos por grupo. A amostra de polissacarídeo foi administrada oralmente em camundongos antes (7 dias) e durante a indução de colite com sulfato de dextran sulfato de sódio (7 dias), seguido por 3 dias adicionais de administração. Alterações no peso corporal e comprimento do cólon foram utilizadas como marcadores para colite, e citocinas pró-inflamatórias e tumores receptor fator 1 (TNFR1), bem como necroptose foram analisadas no cólon de camundongos colite. Os dados obtidos foram analisados por testes Tukey-Kramer e testes padrão t de duas caudas. Resultados: Os resultados indicaram que a amostra de polissacarídeo suprimiu colite em camundongos usando efeitos sobre o peso corporal e o comprimento do cólon como marcadores. Além disso, foi demonstrado que a necrostatina-1, inibidora específica da necroptose, suprimiu a expres são de interleucina (IL)-8, uma quimiocina pró-inflamatória, em células caco-2 induzidas necroptose induzidas por zVAD e TNF-α, uma indicação de que a necroptose pode estar envolvida na expressão de citocinas pro-inflamatórias. Além disso, a amostra de polissacarídeo suprimiu a expressão de citocinas pró-inflamatórias, como o fator de necrose tumoral (TNF)-α, IL-6, IL-1β e interferon (IFN)-γ no cólon dos camundongos. Conclusão: Esses resultados sugeriram que os efeitos supressivos da amostra de polissacarídeo na expressão de citocinas inflamatórias podem contribuir para o seu efeito anti-colite, podendo, portanto, servir como um potente agente terapêutico contra doença inflamatória intestinal.

Taxifolin attenuates inflammation via suppressing MAPK signal pathway in vitro and in silico analysis / 中草药·英文版

Objective: Taxifolin is a natural flavonoid compound that can be isolated from onions, grapes, oranges and grapefruit. It also acts as a medicine food homology with extraordinary antioxidant and anti-inflammatory activity. This study aims to explain the protective effects and potential mechanisms of taxifolin against inflammatory reaction. Methods: Levels of interleukin (IL)-6, IL-1β and intracellular reactive oxygen species (ROS) were assessed in different time after the treatment of taxifolin in RAW264.7 cells induced by lipopolysaccharide (LPS). Subsequently, the mRNA and protein levels of inducible nitric oxide synthase (iNOS), vascular endothelial growth factor (VEGF), cyclooxygenase (COX)-2, tumor necrosis factor (TNF)-α and the phosphorylation expression levels of the MAPK signal pathway were also evaluated. A silico analysis was used to explain the binding situation for the investigation of taxifolin and MAPK signal pathway. And then MAPK inhibitors were used to reveal the expression level of iNOS, VEGF, COX-2 and TNF-α in RAW264.7 cells. Results: It was demonstrated that cell inflammatory damage induced by LPS was significantly alleviated after the treatment of taxifolin. Then, the mRNA and protein levels of iNOS, VEGF, COX-2 and TNF-α were reduced and the phosphorylation expression levels of the MAPK signal pathway were down-regulated remarkably as well. In silico analysis, taxifolin could form a relatively stable combination with MAPK signal pathway. MAPK inhibitors showed increasing or decreasing effect in the mRNA levels of iNOS, VEGF, COX-2 and TNF-α, which suggesting that taxifolin down-regulated iNOS, VEGF, COX-2 and TNF-α expressions were not entirely through the MAPK pathway. Conclusion: This finding demonstrated that taxifolin improved the inflammatory responses that partly involved in the phosphorylation expression level of MAPK signal pathway in RAW264.7 cells exposed to acute stress.

Ovarian hormones influence immune response to Staphylococcus aureus infection

ABSTRACT Objective: Staphylococcus aureus infections remain associated with considerable morbidity and mortality in both hospitals and the community. There is little information regarding the role of ovarian hormones in infections caused by S. aureus. The aim of this study was to evaluate the effects of ovariectomy in the immune response induced by S. aureus. Methods: Female mice BALB/c were ovariectomized (OVX) to significantly reduce the level of ovarian hormones. We also used sham-operated animals. The mice were inoculated intraperitoneally with S. aureus. Blood samples were collected for leukocyte count and bacterial quantification. The uterus and spleen were removed and weighed to calculate the uterine and splenic indexes. Lungs were removed and fractionated for immunohistochemical analysis for macrophage detection (anti-CD68) and relative gene expression of IL-6, IL-1β and TNF-α by RT-PCR. Results: Ovariectomy enlarged spleen size and generally increased circulating lymphocytes. OVX females experienced a continuation of the initial reduction of lymphocytes and a monocyte and neutrophil late response compared to shams (p ≥ 0.05). Moreover, OVX females showed neutropenia after 168 h of infection (p ≥ 0.05). Macrophage response in the lungs were less pronounced in OVX females in the initial hours of infection (p ≥ 0.01). OVX females showed a higher relative gene expression of IL-1β, IL-6 and TNF-α in the lung at the beginning of the infection compared to sham females (p ≥ 0.01). Among the uninfected females, the OVX control females showed a higher expression of IL-6 in the lung compared to the sham control females (p ≥ 0.05). In this model, the lack of ovarian hormones caused a minor increase in circulating leukocytes during the initial stage of infection by S. aureus and increased pulmonary gene expression of IL-1β, IL-6, and TNF-α. Ovariectomy alone enlarged the spleen and increased circulating lymphocytes. Ovarian hormones acted as immunoprotectors against S. aureus infection.

Effect of coinfection with influenza virus and bacteria on host damage / Efecto de la coinfección por virus de la influenza y bacterias en el daño al hospedero

Abstract Background: Influenza virus infection is often complicated by a bacterial infection, with this coinfection causing severe pneumonia. If not timely treated, the disease can cause death. Objective: To demonstrate, in animal models, that coinfection with influenza virus and bacteria that affect the respiratory tract causes multisystemic damage. Method: Six groups of mice were formed: a control group, one infected with the influenza virus, two infected with bacteria: Haemophilus influenzae and Streptococcus pneumoniae, respectively; and two co-infected with influenza virus and Haemophilus influenzae or Streptococcus pneumoniae, respectively. Results: Of the six groups of mice, only the group co-infected with influenza virus and Streptococcus pneumoniae showed damage to thoracic and abdominal organs. A decrease in serum cytokine levels was found in all study groups, which was more pronounced in the co-infected mice. Conclusions: The groups of mice infected with Streptococcus pneumoniae or influenza virus alone showed no damage, which indicates that coexistence of these infections caused the damage in the group of co-infected mice.

Resumen Antecedentes: La infección por el virus de la influenza con frecuencia se complica con una infección bacteriana, coinfección que provoca cuadros graves de neumonía, la cual puede ocasionar la muerte si no es tratada en forma oportuna. Objetivo: Demostrar en modelos animales que la coinfección por el virus de la influenza y bacterias que afectan el tracto respiratorio ocasiona daño multisistémico. Método: Se formaron seis grupos de ratones: un grupo control, uno infectado de virus de la influenza, dos infectados de bacterias: Haemophilus influenzae y Streptococcus pneumoniae, respectivamente; y dos coinfectados de virus de la influenza y Haemophilus influenzae y Streptococcus pneumoniae, respectivamente. Resultados: De los seis grupos de ratones, solo en el grupo coinfectado de virus de la influenza y Streptococcus pneumoniae se observó daño en órganos torácicos y abdominales. En todos los grupos se encontró disminución de los niveles séricos de las citocinas, mayor en los ratones coinfectados. Conclusiones: Los grupos de ratones infectados solo de Streptococcus pneumoniae o el virus de la influenza no presentaron daños, lo cual indica que la coexistencia de estas infecciones fue la que ocasionó el daño en el grupo de ratones coinfectados.

Ophiopogonin D of Ophiopogon japonicus ameliorates renal function by suppressing oxidative stress and inflammatory response in streptozotocin-induced diabetic nephropathy rats

Ophiopogonin D (OP-D) is the principal pharmacologically active ingredient from Ophiopogon japonicas, which has been demonstrated to have numerous pharmacological activities. However, its protective effect against renal damage in streptozotocin (STZ)-induced diabetic nephropathy (DN) rats remains unclear. The present study was performed to investigate the protective effect of OP-D in the STZ-induced DN rat model. DN rats showed renal dysfunction, as evidenced by decreased serum albumin and creatinine clearance, along with increases in serum creatinine, blood urea nitrogen, TGF-β1, and kidney hypertrophy, and these were reversed by OP-D. In addition, STZ induced oxidative damage and inflammatory response in diabetic kidney tissue. These abnormalities were reversed by OP-D treatment. The findings obtained in the present study indicated that OP-D might possess the potential to be a therapeutic agent against DN via inhibiting renal inflammation and oxidative stress.

Efeito da fibrose hepática sobre a severidade da periodontite apical: análise histológica e imunoistoquímica em ratos / Effect of liver fibrosis on the severity of apical periodontitis: histological and immunohistochemical analysis in rats

A inter-relação das alterações sistêmicas com o desenvolvimento e progressão de infecções orais como a periodontite apical tem sido objeto de intensos estudos nos últimos anos. O objetivo deste trabalho foi verificar a influência da fibrose hepática (FH) na severidade da periodontite apical (PA) em ratos Wistar. Quarenta ratos foram divididos em 4 grupos (n=10): Grupo C - ratos controle; Grupo PA - ratos portadores de PA; Grupo FH - ratos portadores de FH; Grupo PA+FH - ratos portadores de PA e FH. A FH foi induzida pelos métodos químico e cirúrgico associados. Foi administrado Tetracloreto de Carbono (CCl4) na dosagem de 0,2ml/100g de peso corporal, 2 vezes por semana, via intraperitoneal, e durante todo o experimento (60 dias). Após 30 dias do início da administração da droga, os animais foram submetidos à cirurgia de ligadura do ducto biliar. Em seguida, as a polpa dentária dos primeiros e segundos molares superiores e inferiores direito foram expostas pelo período de 30 dias para desenvolvimento da PA. Ao final do experimento os animais foram eutanaziados e as maxilas, assim como os fígados, coletados para análise em microscopia de luz. O tecido hepático foi analisado em coloração de hematoxilina e eosina (H&E) e Picrosírius red para comprovar a fibrose hepática e as maxilas processadas para análise histológica, histométrica e imunoistoquímica para IL-1ß, IL-6, e TNFα. Os resultados obtidos foram analisados e comparados por testes estatísticos específicos para cada caso com nível de significância de 5% (p < 0.05). A FH foi confirmada pela análise histológica dos fígados nos grupos FH e PA+FH que apresentaram hepatócitos necrosados, desorganização vascular e intensa deposição de colágeno no parênquima hepático, formando pontes de fibrose. Quanto à periodontite apical observou-se infiltrado inflamatório moderado no grupo PA e intenso no PA+FH (p < 0.05). A análise histométrica mostrou maiores áreas de reabsorção óssea periapical no grupo PA+FH em comparação ao grupo PA (p < 0.05). A análise imunoistoquímica revelou maior imunomarcação para citocinas IL-1ß, IL-6 e TNF-α no grupo PA+FH quando comparado ao grupo PA (p < 0.05). Conclui-se que a FH influencia na severidade da periodontite apical, exacerbando o infiltrado inflamatório, por meio do aumento das citocinas IL-1ß, IL-6 e TNF-α, e aumentando a reabsorção óssea periapical(AU)

The interrelationship between systemic disorders and oral infections such as apical periodontitis has been the subject of intense studies in recent years. The aim of this study was to evaluate the influence of liver fibrosis (LF) on the severity of apical periodontitis (AP) in Wistar rats. Forty rats were divided into 4 groups (n=10): Group C - control rats; AP group - rats with AP; LF Group - rats with LF; AP+LF Group - rats with AP and LF. LF was induced by the association of chemical and surgical methods. Carbon tetrachloride (CCl4) was administered at a dosage of 0.2ml/100g of body weight, twice a week, intraperitoneally, and throughout the experiment (60 days). After 30 days from the beginning of the drug administration, the animals were submitted to bile duct ligation surgery and the dental pulps of the first and second right maxillary and mandibular molars were exposed to induce AP in a period of 30 days. At the end of the experiment, the animals were killed and the jaws, as well as the livers, were collected for analysis under light microscopy. The liver tissue was analyzed in Hematoxilin and Eosin (H&E) and Picrosírius red staining to confirm the liver fibrosis. The jaws were processed for histological, histometric and immunohistochemical analysis for IL1ß, IL-6, and TNF-α. The results obtained were analyzed and compared by specific statistical tests (p < 0.05). LF was confirmed by histological analysis of the liver in the LF and AP+LF groups, which presented necrotic hepatocytes, vascular disorganization and intense collagen deposition in the liver parenchyma, forming fibrosis bridges. A moderate inflammatory infiltrate was observed, in the AP group and intense in the AP+LF (p < 0.05). Histometric analysis showed greater areas of periapical bone resorption in the AP+LF group compared to the AP group (p < 0.05). The immunohistochemical analysis revealed greater immunolabeling for cytokines IL-1ß, IL-6 and TNF-α in the AP+LF group when compared to the AP group (p < 0.05). It is concluded that LF influences on the severity of apical periodontitis, exacerbating the inflammatory infiltrate, by increasing the cytokines IL-1ß, IL-6 and TNF-α, and intensifying the periapical bone resorption(AU)

Effects of leptin-modified human placenta-derived mesenchymal stem cells on angiogenic potential and peripheral inflammation of human umbilical vein endothelial cells (HUVECs) after X-ray radiation / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Combined radiation-wound injury (CRWI) is characterized by blood vessel damage and pro-inflammatory cytokine deficiency. Studies have identified that the direct application of leptin plays a significant role in angiogenesis and inflammation. We established a sustained and stable leptin expression system to study the mechanism. A lentivirus method was employed to explore the angiogenic potential and peripheral inflammation of irradiated human umbilical vein endothelial cells (HUVECs). Leptin was transfected into human placenta-derived mesenchymal stem cells (HPMSCs) with lentiviral vectors. HUVECs were irradiated by X-ray at a single dose of 20 Gy. Transwell migration assay was performed to assess the migration of irradiated HUVECs. Based on the Transwell systems, co-culture systems of HPMSCs and irradiated HUVECs were established. Cell proliferation was measured by cell counting kit-8 (CCK-8) assay. The secretion of pro-inflammatory cytokines (human granulocyte macrophage-colony stimulating factor (GM-CSF), interleukin (IL)-1α, IL-6, and IL-8) was detected by enzyme-linked immunosorbent assay (ELISA). The expression of pro-angiogenic factors (vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF)) mRNA was detected by real-time quantitative polymerase chain reaction (RT-qPCR) assay. Relevant molecules of the nuclear factor-KB (NF-kB) and Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling pathways were detected by western blot assay. Results showed that leptin-modified HPMSCs (HPMSCs/leptin) exhibited better cell proliferation, migration, and angiogenic potential (expressed more VEGF and bFGF). In both the single HPMSCs/leptin and the co-culture systems of HPMSCs/leptin and irradiated HUVECs, the increased secretion of pro-inflammatory cytokines (human GM-CSF, IL-1α, and IL-6) was associated with the interaction of the NF-KB and JAK/STAT signaling pathways. We conclude that HPMSCs/leptin could promote angiogenic potential and peripheral inflammation of HUVECs after X-ray radiation.

Cholinergic anti-inflammatory pathway plays negative regulatory role in early inflammatory and immune responses in septic rats / 南方医科大学学报

OBJECTIVE@#To investigate the role of cholinergic anti-inflammatory pathway (CAP) in neuro-regulation of inflammatory and immune response in the early stage of sepsis.@*METHODS@#Sixty-four SD rats were randomly divided into control Group (=8) with normal feeding without any treatment; sham operation group (=8) with laparotomy but without cecal ligation and puncture (CLP), followed by intraperitoneal injection 50 mg/kg piperacillin 3 times a day for 3 consecutive days; and sepsis group (=48) with CLP-induced sepsis. The rat models of sepsis were randomized into model groups (=16) with intraperitoneal injection of piperacillin (50 mg/kg) and normal saline (1 mL/100 g) for 3 times a day for 3 days; GTS-21 group (=16) with additional intraperitoneal injection of 4 mg/kg GTS-21 (once a day for 3 days); and methyllycaconitine (MLA) group (=16) with intraperitoneal injection of MLA (4.8 mg/kg) in addition to piperacillin (once a day for 3 days). Murine Sepsis Score (MSS) of the rats and short-range HRV analysis were recorded. Three days later, the rats were sacrificed and serum levels of TNF-α, IL-1α, IL-10, IL-6, HMGB1, and sCD14 were measured with ELISA. The percentages of CD4CD25 Treg and TH17 lymphocytes and their ratios were measured using flow cytometry.@*RESULTS@#Compared with the control rats, the septic rats had significantly increased MSS scores and lowered HRV indexes (SDNN, RMSSD, HF, SD1, and SD2; < 0.05); treatment with GTS-21 significantly decreased while MLA increased MSS scores ( < 0.05), but neither of them obviously affected HRV of the rats. Serum levels TNF-α, IL-1α, IL-10, IL-6, HMGB1, and sCD14 and the percentages of CD4CD25 Treg and TH17-positive lymphocytes were significantly higher and Treg/TH17 ratio was significantly lower in the septic rats compared with those in the control group ( < 0.05); treatment with GTS-21 significantly decreased the levels of serum levels of TNF-α, IL-1α, IL-6, HMGB1, and sCD14 and TH17 lymphocyte percentage ( < 0.05), whereas MLA treatment significantly increased serum levels of TNF-α, IL-1α, IL-10, IL-6, HMGB1, and sCD14 and the percentages of CD4 CD25 Treg and TH17-positive lymphocytes and decreased Treg/TH17 ratio in the septic rats ( < 0.05).@*CONCLUSIONS@#CAP plays negative regulatory role in early inflammatory and immune response to sepsis, and some of the HRV indicators can well reflect the regulatory effect of CAP on inflammation and immunity in the septic rats.

Attenuation of lipopolysaccharide-induced neuroinflammatory events in BV-2 microglial cells by Moringa oleifera leaf extract

Objective: To determine the anti-neuroinflammatory activity of Moringa oleifera leaf extract (MLE) under lipopolysaccharide stimulation of mouse murine microglia BV2 cells in vitro. Methods: The cytotoxicity effect of MLE was investigated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide assay. The inflammatory response of BV-2 cells were induced with lipopolysaccharide. The generation of nitric oxide levels was determined by using Griess assay and the level of pro-inflammatory cytokines (IL-1β, IL-6 and TNF-α) was evaluated by ELISA kit. The expression of iNOS, COX-2 as well as IκB-α was carried out by immunoblot analysis. Results: MLE reduced the nitric oxide production in concentration-dependent manner, and maintained the viability of BV-2 microglial cells which indicated absence of toxicity. In addition, MLE repressed the activation of nuclear factor kappa B by arresting the deterioration of IκB-α, consequently resulted in suppression of cytokines expression such as COX-2 and iNOS. Conclusions: MLE inhibitory activities are associated with the inhibition of nuclear factor kappa B transcriptional activity in BV2 microglial cells. Thus MLE may offer a substantial treatment for neuroinflammatory diseases.

Adipose tissue is less responsive to food restriction anti-inflammatory effects than liver, muscle, and brain in mice

High caloric intake promotes chronic inflammation, insulin resistance, and chronic diseases such as type-2 diabetes, which may be prevented by food restriction (FR). The effect of FR on expression of pro-inflammatory and anti-inflammatory genes in adipose tissue, liver, muscle, and brain was compared. Male Swiss mice were submitted to FR (FR group) or had free access to food (control group) during 56 days. The liver, gastrocnemius muscle, brain, and epididymal white adipose tissue (WAT) were collected for analysis of gene expressions. FR attenuated inflammation in the liver, brain, and gastrocnemius muscle but did not markedly change inflammatory gene expression in epididymal WAT. We concluded that adipose tissue was less responsive to FR in terms of gene expression of pro-inflammatory and anti-inflammatory genes.

Moxibustion promoted transformation of inflammatory phase to facilitate wound healing in rats with full-thickness cutaneous wounds / 针刺研究

OBJECTIVE: Our previous study demonstrated that moxibustion could promote skin wound healing in rats with full-thickness cutaneous wounds. The present study was designed to observe the effect of moxibustion on the levels of pro-inflammatory and anti-inflammatory cytokines, and proliferation of vascular endothelial cells, so as to explore its mechanisms underlying facilitating wound-healing. METHODS: A total of 84 adult SD male rats were randomly assigned to normal (n＝6), model(n＝39)and moxibustion(n＝39)groups. The skin wound model was established by removal of a piece of full-thickness skin from the median line of the rats' back (about 2 cm below the shoulder blade). Moxibustion was applied to the surrounding area of the focus for 25 min, once daily for 6 days. The blood samples were collected at day 1, 2, 3 and 5 after modeling for assaying serum contents of IL-1α， IL-1β， IL-6, IL-4, IL-10, IL-13, granulocyte-colony stimulating factor (G-CSF), granulocyte macrophagecolony stimulating factor (GM-CSF, an inflammatory mediator), macrophage inhibitory protein-1α (MIP-1α) and vascular endothelial growth factor (VEGF) with liquid chip methods, and the topical wound tissues were collected after trans-cardiac perfusion with 4% paraforma-ldehyde solution for detecting the expression of CD31 proteins by using immunofluorescence staining. RESULTS: After modeling and in comparison with the model group, the contents of serum IL-1α and IL-6 at the 1st day were significantly increased (P<0.05), but notably decreased at the 3rd day after modeling in the moxibustion group (P<0.05). The contents of serum IL-1β on day 1 and 2 were significantly higher in the moxibustion group than those of the model group (P<0.05). After moxibustion, the contents of serum IL-4 and IL-10 were significantly up-regulated on day 1 after modeling (P<0.05), and obviously down-regulated from the 3rd day on (P<0.05). The contents of serum MIP-1α on the 2nd day and serum VEGF on day 1 and 2, and wound-skin VEGF on day 5 were obviously up-regulated in the moxibustion group in comparison with the model group (P<0.05, P<0.01)．. CONCLUSION: Moxibustion promotes the inflammatory response by regulating the levels of both pro-inflammatory and anti-inflammatory cytokines in the early phase of skin wounds in rats, which may be in favor of the transformation from the inflammatory phase to the proliferation phase in advance, promoting wound healing at last.

Effect and Mechanisms of Lycii Fructus on Hepatoma HepG2 Cell Line-induced Cachexia in Mice / 中国实验方剂学杂志

Objective: To investigate the inhibitory effect and mechanism of the extracts from fresh Lycii Fructus (LBL) on hepatoma HepG2 cell-induced cachexia in mouse.Method: The human hepatoma cell line HepG2 was injected into BALB/C mice to establish the cachexia model.Then the LBL was fed to the models respectively in low dose (5 mg·kg-1) or high dose (25 mg·kg-1).After 28 days of continuous feeding,the mice's body weight was detected.The expression levels of creatine kinase (CK),interleukin-1β (IL-1β),interleukin-6(IL-6) and tumor necrosis factor-α(TNF-α) were evaluated by enzyme linked immunosorbent assay (ELISA).The muscle degradation and the expression of p38 mitogen-activated protein kinase (p38 MAPK) were detected by immunohistochemistry staining.The expression levels of phospho-p38 mitogen-activated protein kinase (p-p38 MAPK) and nuclear factor-κB (NF-κB) were determined by Western blot.Result: Compared with cachexia group,the loss of body weight and muscle decomposition were significantly inhibited both in the low dose LBL group and high dose LBL group (P0.05,PPβ,IL-6 and TNF-α in both the low-dose LBL group and the high-dose LBL group (PPPκB were inhibited both in the low-dose LBL group and the high-dose LBL group (PConclusion: LBL could inhibit the cachexia induced by hepatoma HepG2 cell line in mouse,suggesting LBL could reduce major cytokine and plasma inflammatory factors through p-p38 MAPK pathway.

Blockade of Endogenous Angiotensin-(1-7) in Hypothalamic Paraventricular Nucleus Attenuates High Salt-Induced Sympathoexcitation and Hypertension / 神经科学通报·英文版

Angiotensin (Ang)-(1-7) is an important biologically-active peptide of the renin-angiotensin system. This study was designed to determine whether inhibition of Ang-(1-7) in the hypothalamic paraventricular nucleus (PVN) attenuates sympathetic activity and elevates blood pressure by modulating pro-inflammatory cytokines (PICs) and oxidative stress in the PVN in salt-induced hypertension. Rats were fed either a high-salt (8% NaCl) or a normal salt diet (0.3% NaCl) for 10 weeks, followed by bilateral microinjections of the Ang-(1-7) antagonist A-779 or vehicle into the PVN. We found that the mean arterial pressure (MAP), renal sympathetic nerve activity (RSNA), and plasma norepinephrine (NE) were significantly increased in salt-induced hypertensive rats. The high-salt diet also resulted in higher levels of the PICs interleukin-6, interleukin-1beta, tumor necrosis factor alpha, and monocyte chemotactic protein-1, as well as higher gp91 expression and superoxide production in the PVN. Microinjection of A-779 (3 nmol/50 nL) into the bilateral PVN of hypertensive rats not only attenuated MAP, RSNA, and NE, but also decreased the PICs and oxidative stress in the PVN. These results suggest that the increased MAP and sympathetic activity in salt-induced hypertension can be suppressed by blockade of endogenous Ang-(1-7) in the PVN, through modulation of PICs and oxidative stress.

Attenuation of lipopolysaccharide-induced neuroinflammatory events in BV-2 microglial cells by Moringa oleifera leaf extract

Objective: To determine the anti-neuroinflammatory activity of Moringa oleifera leaf extract (MLE) under lipopolysaccharide stimulation of mouse murine microglia BV2 cells in vitro. Methods: The cytotoxicity effect of MLE was investigated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5- diphenyl-tetrazolium bromide assay. The inflammatory response of BV-2 cells were induced with lipopolysaccharide. The generation of nitric oxide levels was determined by using Griess assay and the level of pro-inflammatory cytokines (IL-1β, IL-6 and TNF-α) was evaluated by ELISA kit. The expression of iNOS, COX-2 as well as IκB-α was carried out by immunoblot analysis. Results: MLE reduced the nitric oxide production in concentration-dependent manner, and maintained the viability of BV-2 microglial cells which indicated absence of toxicity. In addition, MLE repressed the activation of nuclear factor kappa B by arresting the deterioration of IκB-α, consequently resulted in suppression of cytokines expression such as COX-2 and iNOS. Conclusions: MLE inhibitory activities are associated with the inhibition of nuclear factor kappa B transcriptional activity in BV2 microglial cells. Thus MLE may offer a substantial treatment for neuroinflammatory diseases.