Structure-activity relationship of novel series of 1,5-disubstituted tetrazoles as cyclooxygenase-2 inhibitors: Design, synthesis, bioassay screening and molecular docking studies.

A novel class of modified 1,5-disubstituted tetrazoles was designed and synthesized, their biological activity as cyclooxygenases inhibitors was screened, and their molecular docking studies were performed. The structural modifications of the first category included the 4-methylsulfonyl phenyl at C-1 of the central moiety and the linkers (-OH, -CH2OH, -CH2CH2OH) with different lengths at the para position of the N-1 phenyl group. For the second category, the 4-methylsulfonyl phenyl group at C-1 was replaced with 4-aminosulfonyl phenyl. While for the third category, a methylene unit was inserted between the C-1 of the tetrazole central ring and the 4-(methylsulfonyl)phenyl group, keeping the same linkers of various extensions at the para position of the N-1 phenyl group. Among the screened compounds, tetrazole 4i showed the best inhibition potency and selectivity values for both COX-2 enzyme (IC50=3µM, SI>67) and COX-1 isoenzyme (IC50>200µM). Compounds 4e, 4h, and 4i, which have the highest inhibition potency toward COX-2 were selected for the molecular docking studies to verify their inhibition and selectivity for COX-2 over COX-1 with their modified structure. The obtained theoretical studies are in agreement with the in vitro bioassay screening results, which supports the importance of the structural modifications for our studied compounds.

Pomegranate and licorice juices modulate metformin pharmacokinetics in rats.

OBJECTIVES: Food or drinks may significantly alter the pharmacokinetics and pharmacodynamics of drugs which may lead to adverse events. A drug such as metformin is widely used to regulate plasma glucose level and pomegranate and licorice have been identified to help in type-2 diabetes management. However, the interactions of the latter on metformin pharmacokinetics were not studied. Therefore, we aimed here to study the impact of pomegranate and licorice on metformin pharmacokinetics in rats. METHODS: Juices were given to rats for two days and half an hour before metformin (20 mg/kg) oral administration. Blood samples, then, were collected at different time intervals, processed and analyzed using validated reliable HPLC method. Plasma profile and pharmacokinetic parameters were calculated for each group. RESULTS AND CONCLUSION: Pre-administration of pomegranate significantly reduced metformin maximum plasma concentration from 1410 to 1031 ng/ml. On the other hand, pre-administration of licorice significantly delayed metformin reaching its maximum plasma concentration. In conclusion, pre-administration of pomegranate may potentially reduce efficacy of metformin while licorice might delay metformin action. Thus, both juices should be cautiously administrated with metformin, the mainstay drug for type-2 diabetes mellitus management.

Eriobotrya japonica Water Extract Characterization: An Inducer of Interferon-Gamma Production Mainly by the JAK-STAT Pathway.

Eriobotrya japonica (Thunb.) Lindl. (Loquat) (EJ) has been used as a medicinal plant to treat chronic bronchitis, coughs, phlegm, high fever and gastro-enteric disorders. Since the traditional use of EJ is related to modulating inflammation processes, our earlier studies on EJ leaves were performed on the water extract to investigate specific cytokines' modulation. These earlier studies, however, have shown that EJ leaf water extract (WE) and the water phase (WP) induce cytokines' production in in vitro and in vivo models. Therefore, the aim of this study was to specify the group(s) of compounds in EJ leaves that have this immunomodulatory activity and their mechanism of action. WE was obtained from boiling the leaves followed by butanol extraction, yielding a butanol-water phase (WP). WP was then subjected to methanol:acetone fractionation, yielding upper (MAU) and lower (MAL) phases. For further fractionation, MAU was subjected to column chromatography followed by elution with ethanol:water (EW), methanol:ethanol (ME) and, lastly, acetone:water (AW), respectively, to reveal three sub-fractions; MAU-EW, MAU-ME and MAU-AW. MAU-AW significantly increased IFN-γ production from unstimulated and stimulated mouse spleen cells, as well as CD3+ T cells and natural killer cells. Furthermore, the fold increase of IFN-γ production by MAU-AW was concentration dependent, higher than the parent extract or any of the other sub-fractions, and such an IFN-γ increase was reversed by two JAK-STAT inhibitors. In addition, MALDI-TOF-MS analysis of the extracts and sub-fractions showed compounds with molecular weights of >500 Daltons. The MAU-AW sub-fraction contained more polar compounds, such as flavonol and caffeic glycosides. In conclusion, these polar compounds in the EJ extract are responsible for inducing IFN-γ production. Further chemical elucidation is warranted to lead to a specific IFN-γ inducer and an immunomodulator in polarizing immune cells and balancing immune responses in certain diseases.

IFN-γ, IL-17 and TGF-ß involvement in shaping the tumor microenvironment: The significance of modulating such cytokines in treating malignant solid tumors.

Multiple innate and adaptive immune effector cells and molecules partake in the recognition and destruction of cancer cells to protect against growing tumors, a concept that is known as cancer immunosurveillance. Unfortunately, cancer cells are capable of avoiding this process by immunoselection of poorly immunogenic tumor cells variants along with subversion of the immune system and thus shaping both the tumor and its microenvironment. Cytokines represent part of the complex pattern of the immune response which can assist the development of cancer as well as to eliminate it. Simultaneously, a large number of cytokines may be involved in the complex interactions between host and tumor cells where this dynamic cross-talk, between tumors and the immune system, can either regulate tumor growth or tumor growth, invasion and metastasis take place. In this review, we are stressing on the interface between infiltrated immune cells and tumor cells with the emphasis on the bidirectional activities of specific cytokines: IFN-γ, TGF-ß and IL-17 within the tumor microenvironment and their role in shaping it. In addition, the significance of modulating such cytokines in favor of anti-tumor response is discussed and merits the use of mixture of targeted modulators to overcome the network complexity of cytokines in the tumor microenvironment.

Plasma concentrations of 25-hydroxyvitamin D among Jordanians: Effect of biological and habitual factors on vitamin D status.

BACKGROUND: Vitamin D is cutaneously synthesized following sun exposure (vitamin D3) as well as it is derived from dietary intake (vitamin D3 and D2). Vitamin D2 and D3 are metabolized in the liver to 25-hydroxyvitamin D (25(OH)D). This metabolite is considered the functional indicator of vitamin D stores in humans. Since Jordan latitude is 31°N, cutaneous synthesis of vitamin D3 should be sufficient all year round. However, many indications reveal that it is not the case. Thus, this study was conducted to determine the 25(OH)D status among Jordanians. METHODS: Three hundred healthy volunteers were enrolled in a cross sectional study; 201 females and 99 males. 25(OH)D and calcium concentrations were measured by enzyme linked immunosorbent assay and spectroscopy techniques, respectively. All participants filled a study questionnaire that covered age, sex, height, weight, diet, and dress style for females. Females were divided according to their dress style: Western style, Hijab (all body parts are covered except the face and hands), and Niqab (all body parts are covered including face and hands). RESULTS: The average plasma 25(OH)D levels in males and females were 44.5 ± 10.0 nmol/l and 31.1 ± 12.0 nmol/l, respectively. However, when female 25(OH)D levels were categorized according to dress styles, the averages became 40.3, 31.3 and 28.5 nmol/l for the Western style, Hijab and Niqab groups, respectively. These 25(OH)D levels were significantly less than those of males (p < 0.05, 0.001, 0.001, respectively). In addition, the plasma 25(OH)D levels of the Western style group was significantly higher than those of Hijab and Niqab groups (p < 0.001). Furthermore, dairy consumption in males was a positive significant factor in vitamin D status. Even though calcium concentrations were within the reference range, the Hijab and Niqab-dressed females have significantly less plasma calcium levels than males (p < 0.01). CONCLUSIONS: Very low plasma 25(OH)D levels in females wearing Hijab or Niqab are highly attributed to low sunlight or UVB exposure. In addition, most of males (76%) and Western style dressed females (90%) have 25(OH)D concentrations below the international recommended values (50 nmol/l), suggesting that although sun exposure should be enough, other factors do play a role in these low concentrations. These findings emphasize the importance of vitamin D supplementation especially among conservatively dressed females, and determining if single nucleotide polymorphisms of the genes involved in vitamin D metabolism do exist among Jordanians.

Eriobotrya japonica hydrophilic extract modulates cytokines in normal tissues, in the tumor of Meth-A-fibrosarcoma bearing mice, and enhances their survival time.

BACKGROUND: Cytokines play a key role in the immune response to developing tumors, and therefore modulating their levels and actions provides innovative strategies for enhancing the activity of antigen presenting cells and polarizing towards T helper 1 type response within tumor microenvironment. One of these approaches could be the employment of plant extracts that have cytokine immunomodulation capabilities. Previously, we have shown that the Eriobotrya japonica hydrophilic extract (EJHE) induces proinflammatory cytokines in vitro and in vivo. METHODS: The present study explored the in vivo immunomodulatory effect on interferon-gamma (IFN-γ), interleukin-17 (IL-17), and transforming growth factor-beta 1 (TGF-ß1) evoked by two water-extracts prepared from EJ leaves in the tissues of normal and Meth-A-fibrosarcoma bearing mice. RESULTS: Intraperitoneal (i.p.) administration of 10 µg of EJHE and EJHE-water residue (WR), prepared from butanol extraction, increased significantly IFN-γ production in the spleen (p < 0.01) and lung (p < 0.03) tissues at 6-48 hours and suppressed significantly TGF-ß1 production levels (p < 0.001) in the spleen for as long as 48 hours. The latter responses, however, were not seen in Meth-A fibrosarcoma-bearing mice. On the contrary, triple i.p. injections, 24 hours apart; of 10 µg EJHE increased significantly IFN-γ production in the spleen (p < 0.02) while only EJHE-WR increased significantly IFN-γ, TGF-ß1 and IL-17 (p < 0.03 - 0.005) production within the tumor microenvironment of Meth-A fibrosarcoma. In addition, the present work revealed a significant prolongation of survival time (median survival time 72 days vs. 27 days of control, p < 0.007) of mice inoculated i.p. with Meth-A cells followed by three times/week for eight weeks of i.p. administration of EJHE-WR. The latter prolonged survival effect was not seen with EJHE. CONCLUSIONS: The therapeutic value of EJHE-WR as an anticancer agent merits further investigation of understanding the effect of immunomodulators' constituents on the cellular components of the tissue microenvironment. This can lead to the development of improved strategies for cancer treatment and thus opening up a new frontier for future studies.

Dopamine selectively modulates lipopolysaccharide-induced TNF-alpha, IFN-gamma and IL-10 within mice tissues.

OBJECTIVES: Dopamine (DA) administration in sepsis is used to modulate the hypotensive condition and to normalize the blood vessels perfusion. However, whether this administration of DA has an effect on the release of cytokines in vivo deserves investigation. METHODS AND RESULTS: Pre-exposure of DA (1 µg/ml) to whole blood enhanced IL-10 (30%) production level following LPS stimulation. This IL-10 enhancement became statistically significant (p<0.001) upon the addition of D2-DA receptor (DAR) antagonists, Clozapine or Haloperidol. Furthermore, systemic administration of DA (0.5-50 mg/kg) in mice suppressed significantly LPS-induced TNF-α levels in blood, liver, spleen, brain, and lungs; IL-10 levels in blood, brain and liver; and IFN-γ levels in blood, liver, brain, and lungs. On the other hand, DA enhanced significantly LPS-induced IL-10 production in the lungs and spleen, and IFN-γ levels in the spleen. Administration of Clozapine (54 mg/kg) or Haloperidol (62 mg/kg) with LPS (1 µg) and DA (5 mg/kg) reversed DA suppressive effects on LPS-induced cytokines in blood, IFN-γ in brain and lungs, and enhanced significantly LPS-induced IL-10 production in blood, spleen, liver, and lungs. CONCLUSIONS: These results indicate that DA modulatory effect on LPS-induced blood cytokines-producing cells is mediated mainly by D2-DAR (D2/ D3/D4) through enhancing immune cells migration and extravasation into tissues. Furthermore, DA selectivity on cytokines modulation is tissue specific, mediated by the type of DAR expressed and on the immune cells lodged in each tissue.

Age of Natural Menopause Among Jordanian Women and Factors Related to Premature and Early Menopause.

OBJECTIVE: The aim of this study was to assess factors related to the onset of premature/early natural menopause among Jordanian women. METHODS: A cross-sectional study was conducted in early 2016. Subjects were enrolled based on random drop-off technique to the Obstetrics and Gynecology clinics at the Jordan University Hospital. Women 18 years of age and above were initially eligible to enroll, and women who had surgically induced menopause or specific disease were excluded from the analysis. Relevant data were collected using a questionnaire that included 30 questions. The following variables were collected: socio-demographic, body mass index, chronic conditions, diseases, reproductive characteristics, and health status. Hormone indicators of menopause were tested by measuring estrogen (E2) and follicle-stimulating hormone (FSH) levels. Age at natural menopause (ANM) was self-reported retrospectively and considered an independent variable against BMI, smoking, hormone therapy, and concomitant diseases. Association analysis and multinomial logistic regression were used to examine the associated factors of ANM with adjusted odds ratios (ORs), and their 95% confidence intervals (CIs) were reported. RESULTS: A total of 409 women were included in the analysis, aged between 20-75 years. The mean ANM in our sample was 48.5±5.0, with 2.7% of the women experienced premature menopause (ANM <40) and 7.8% early menopause (ANM 40-44). Within the menopause women (n=242), the percentage of women who had premature menopause was 4.5%, 13.6% with early menopause, and 21.1% with late menopause (ANM >52). Smoking was the major risk factor for premature/early menopausal age among Jordanian women with an OR of 2.46 (95% CI: 1.08-5.59, p<0.05). On the other hand, women with occasional arthritis symptoms and diseases such as hypertension, diabetes, dyslipidemia, and their combination were associated with average (45-52 years) or late menopause (>52 years). CONCLUSION: Smoking is the main contributor of premature/early menopause in Jordanian women. Increased awareness and public health policy about the adverse effects of smoking on women's reproductive health are needed.

The Prevalence of Overweight and Obesity Among Women in Jordan: A Risk Factor for Developing Chronic Diseases.

OBJECTIVE: The study aimed to investigate the prevalence of obesity among Jordanian women and its association with a wide range of chronic diseases. METHODS: Subjects were enrolled in the present cross-sectional study based on a random drop-off technique at the Obstetrics and Gynecology clinics at Jordan University Hospital. Initially, any female 18 years of age and older was asked to enroll in the study. Relevant data were gathered using a questionnaire composed of 30 questions, and body mass index (BMI) was determined from each participant's weight and height. The following variables were collected: socio-demographic, chronic diseases, and health status. Each variable's frequencies were reported, and the 95% confidence interval (95% CI) for each variable was calculated. For association analysis, Chi-square analysis was performed with an odds ratio (OR) and 95% CI. Multinomial logistic regression analysis was applied to a combination of independent variables and a dependent condition with covariate factors. RESULTS: The age-standardized prevalence of overweight/obese Jordanian women was 70.6% (95% CI 66.0-74.8%). On the other hand, the age-standardized prevalence of only obese women was 36.4 (95% Cl 31.9-41.2%). Furthermore, the association between age and overweight/obesity was significant (p<0.0001). The percentage of overweight and obesity started to be significant in the 30-39 year age group. Moreover, the OR for obesity ranged from 2.7 to 7.0 (p<0.05-0.01) for those women with only elementary education. Besides, high parity was significantly associated with obesity and elementary education. For chronic conditions, the percentages of hypertension, diabetes, hypertriglyceridemia, osteoporosis, and rheumatoid arthritis were significantly correlated with increased BMI in Jordanian women. With age adjustment, however, only hypertension was associated with obese level 3 with OR of 7.2 and 95% CI of 2.1-25.1 (p<0.01). CONCLUSION: There is a high prevalence of overweight/obesity among women in Jordan, which was related to high parity and low education level. This high prevalence of obesity increased the incidence of chronic diseases, such as hypertension. Therefore, community-based multiple strategies are required to combat obesity in Jordanian women.

A bioactive prodelphinidin from Mangifera indica leaf extract.

A new trimeric proanthocyanidin, epigallocatechin-3-O-gallat-(4beta-->8)-epigallocatechin-(4beta-->8)-catechin (1), was isolated together with three known flavan-3-ols, catechin (2), epicatechin (3), and epigallocatechin (4), and three dimeric proanthocyanidins, 5-7, from the air-dried leaves of Mangifera indica. Their chemical structures were determined on the basis of 1D- and 2D-NMR spectra (HSQC, HMBC) of their peracetylated derivatives, MALDI-TOF-mass spectra, and by acid-catalyzed degradation with phloroglucinol. The isolated compounds 1-7 were in vitro tested for their inhibitory activities against COX-1 and COX-2. Compound 1 was found to have a potent inhibitory effect on COX-2, while compounds 1 and 5-7 exhibited moderate inhibition against COX-1.

Pleiotropic Anticancer Properties of Scorpion Venom Peptides: Rhopalurus princeps Venom as an Anticancer Agent.

To date, the success of conventional chemotherapy, radiotherapy, and targeted biological therapies in cancer treatment is not satisfactory. The main reasons for such outcomes rely on low target selectivity, primarily in chemo- and radiotherapy, ineffectiveness to metastatic disease, drug resistance, and severe side effects. Although immune checkpoint inhibitors may offer better clinical promise, success is still limited. Since cancer is a complex systemic disease, the need for new therapeutic modalities that can target or block several steps of cancer cell characteristics, modulate or repolarize immune cells, and are less toxic to healthy tissues is essential. Of these promising therapeutic modalities are pleiotropic natural products in which scorpion venom (SV) is an excellent example. SV consists of complex bioactive peptides that are disulfide-rich of different peptides' length, potent, stable, and exerts various multi-pharmacological actions. SV peptides also contain ion channel inhibitors. These ion channels are dysregulated and overexpressed in cancer cells, and play essential roles in cancer development and invasion, as well as depolarizing immune cells. Furthermore, SV has been found to induce cancer cell apoptosis, and inhibit cancer cells proliferation, invasion, metastasis, and angiogenesis. In the current review, we are presenting data that show the pleiotropic effect of SV against different types of human cancer as well as revealing one potential anticancer agent, Rhopalurus princeps venom. Furthermore, we are addressing what is needed to be done to translate these potential cancer therapeutics to the clinic.

Pseudomonas aeruginosa PAO1 resistance to Zinc pyrithione: phenotypic changes suggest the involvement of efflux pumps.

The aim of this study is to investigate the involvement of an efflux pump in the development of Pseudomonas aeruginosa resistance to zinc pyrithione (ZnPT). In the presence of efflux inhibitor carbonyl cyanide m-chlorophenyl-hydrazone (CCCP), the minimum inhibitory concentration of ZnPT for P. aeruginosa resistant cells is reduced significantly (p < 0.05). In addition, the concentration of ZnPT excluded by the resistant bacteria was reduced significantly (p < 0.01). However, the above reductions did not reach the levels measured for P. aeruginosa PAO1 sensitive strain. Furthermore, such changes in P. aeruginosa resistant cells were correlated with the overexpression of outer membrane proteins, reduced sensitivity toward imipenem (p < 0.01) and increased sensitivity toward sulphatriad and chloramphenicol (p < 0.05). In a continuation to a previous study, we conclude that P. aeruginosa resistance to ZnPT is multifactorial and involves induced efflux systems.

Epicatechin suppresses IL-6, IL-8 and enhances IL-10 production with NF-kappaB nuclear translocation in whole blood stimulated system.

OBJECTIVES: Cytokines induce the balance between inflammatory versus regulatory or antibody mediated reactions. So modulating the release of cytokines or inducing them by immunomodulating agents is an attractive mode for treating or help in treating several diseases such as autoimmune diseases. Eriobotrya japonica is a plant that is traditionally thought to have anti-inflammatory activities. Several compounds were isolated from the plant and showed distinctive biological effects. The purpose of this study was to determine the effects of epicatechin (EC) isolated from Eriobotrya Japonica on IL-6, IL-8, and IL-10 productions in whole blood stimulated with phytohemagglutinin (PHA)+lipopolysaccharide (LPS), and to examine if these cytokines are modulated through NFkappaB pathway. METHODS: Sixteen healthy males and females volunteered in the study. Blood samples were drawn, diluted, and cultured for 24 h with different concentrations of EC and then PHA+LPS was added for another 24 h. The supernatant, then, was harvested and assayed for cytokines. In addition, mixing studies of EC and hydrocortisone were performed to examine the cytoplasmic and nuclear fractions of NFkB levels in association with cytokine production levels. RESULTS: Increasing concentrations of EC (1-100 microg/ml) in PHA+LPS stimulated whole blood cells culture suppressed significantly (p<0.001) the production of IL-6 and IL-8. Moreover, increasing concentrations of EC modulated significantly the production of IL-10, as there was a significant increase in IL-10 level at 0.1, 1.0, and 10 microg/ml (p=0.058-0.004), while a significant decrease at a concentration of 100 microg/ml EC (p=0.037) was observed. In addition, an additive effect between EC and hydrocortisone (HC, 100 nmol/l) was seen in the production of IL-10, as there was a significant increase in IL-10 level (32%) compared with 27% for EC (10 microg/ml) and 19% HC. Furthermore, a significant decrease in cytoplasmic fractions of NFkappaB p65 level was found in samples containing EC 1, 10 microg/ml, but not in 100 microg/ml, when compared with control (p<0.03). These latter changes were accompanied with a 29%, 67%, and 98% increase, respectively, of NFkappaB p65 in nuclear fractions compared to 24% reduction of NFkappaB p65 level in HC culture (p<0.03). CONCLUSIONS: These results indicate that EC suppresses the production of pro-inflammatory cytokines, IL6 and IL-8, enhances the production of anti-inflammatory cytokine, IL-10, and stimulates NFkappaB p65 translocation to nucleus in PHA+LPS stimulated whole blood culture.

A Functional Food Mixture "Protector" Reinforces the Protective Immune Parameters against Viral Flu Infection in Mice.

Background: Viral influenza infection causes serious health issues especially when an outbreak occurs. Although influenza virus vaccines are available and each year manufactures modify the vaccine depending on the expected mutated strain, it is still far from satisfactory, mainly in young children and older adults. Therefore, a product that can support and shape the immune system to protect against viral flu infections is highly essential. Methods: A functional food water-soluble mixture of pomegranate, red grape, dates, olive fruit, figs, and ginger extracts, termed herein “Protector”, was prepared and tested in stimulating/modulating the production of specific cytokines, and hemagglutinin inhibition (HAI) antibodies following viral flu vaccination in mice. Results: A single intraperitoneal or multiple oral administration for 1⁻7 days of “Protector” significantly increased the production of interferon (IFN)-γ and interleukin (IL)-12 in blood, spleen, and lungs of mice. When “Protector” was orally administered for one week following a single vaccine injection (primary immunization) or for two weeks (one week apart) following double vaccine injections (secondary immunization), mice significantly produced higher titers of HAI antibodies. This increase in HAI antibodies was associated with Pillow-inducing significant and different changes in vaccine-induced IFN-γ, IL-12, IL-6 and IL-22 following primary and secondary immunizations. Conclusions: “Protector” administration reinforces the protective immune parameters against viral flu infection. Therefore, after performing preclinical toxicology studies and ensuring its safety, “Protector” should be considered a potential product to be tested in clinical trials to conclude its efficacy in reducing the devastating effects of flu infection in humans and its outbreaks.

The differential effect of Eriobotrya japonica hydrophilic leaf extract on cytokines production and modulation.

Stimulating or modulating the release of cytokines by immunomodulators or immunostimulating agents is an attractive mode for treating several diseases such as viral infections. For instance, patients with viral infections may be in need of increasing or inducing T helper 1 (Th1) or proinflammatory cytokines, which ultimately activate T cytotoxic and Natural killer lymphocytes to kill virally infected cells. Of these agents, we found that Eriobotrya japonica hydrophilic leaf extract (EJHE) can induce and modulate cytokines in dose-dependent manner. Twenty-four hour exposure of increasing concentrations of EJHE increased significantly (p<0.001) the production of IFN-gamma and TNF-alpha, from PHA+LPS-stimulated whole blood. However, the production of IFN-gamma and TNF-alpha plateaued at high EJHE concentrations (10-100 microg/ml). No significant changes in the production of IL-10 were seen. In addition, EJHE at 1 and 10 microg/ml reversed significantly (p < 0.01) the inhibitory effect of hydrocortisone on the IL-12 p70, IFN-gamma and TNF-alpha production from PHAS+LPS stimulated whole blood. Without PHA and LPS, EJHE was found to induce significantly (p<0.001) IFN-gamma, IL-12 p70, TNF-alpha, and IL-10 from whole blood culture in concentration dependent manner. The maximum induction of IFN-gamma, IL-12 p70, and TNF-alpha by EJHE was at 1 and 10 microg/ml. On the other hand, IL-10 induction kept increasing even at the highest concentration used (100 microg/ml) of EJHE. Furthermore, intra-peritoneal injection of EJHE into mice increased significantly serum cytokines level mainly at 10 and 100 microg/ml. Two-hour post i.p. injection, EJHE increased serum IFN-gamma, TNF-alpha, and IL-10 to approximately 750, 1000, and 250 pg/ml, respectively. However, 24h post i.p. injection, the levels of TNF-alpha, and IL-10 were similar to basal levels but IFN-gamma levels were 200 pg/ml. These results indicate that EJHE induces proinflammatory and Th1 cytokines in concentration dependent manner and the effect of this induction should be studied further in viral models to check the efficacy of such cytokine induction.

Glucosamine modulates propranolol pharmacokinetics via intestinal permeability in rats.

Propranolol (PROP) undergoes extensive first-pass metabolism by the liver resulting in a relatively low bioavailability (13-23%); thus, multiple oral doses are required to achieve therapeutic effect. Since some studies have reported that glucosamine (GlcN) can increase the bioavailability of some drugs, therefore, it is aimed to study whether GlcN can change the pharmacokinetic parameters of PROP, thus modulating its bioavailability. When PROP was orally co-administered with GlcN (200mg/kg) to rats, PROP area under curve (AUC) and maximum concentration (Cmax) were significantly decreased by 43% (p<0.01) and 33% (p<0.05), respectively. In line with the in vivo results, in silico simulations confirmed that GlcN decreased rat intestinal effective permeability (Peff) and increased PROP clearance by 50%. However, in situ single pass intestinal perfusion (SPIP) experiments showed that GlcN significantly increased PROP serum levels (p<0.05). Furthermore, GlcN decreased PROP disposition/distribution into cultured hepatocytes in concentration dependent manner. Such change in the interaction pattern between GlcN and PROP might be attributed to the environment of the physiological buffer used in the in vitro experiments (pH7.2) versus the oral administration and thus, enhanced PROP permeability. Nevertheless, such enhancement was not detected when everted gut sacks were incubated with both drugs at the same pH in vitro. In conclusion, GlcN decreased PROP serum levels in rats in a dose-dependent manner. Such interaction might be attributed to decreased intestinal permeability and enhanced clearance of PROP in the presence of GlcN. Further investigations are still warranted to explain the in vitro inhibitory action of GlcN on PROP hepatocytes disposition and the involvement of GlcN in the intestinal and hepatic metabolizing enzymes of PROP at different experimental conditions.

Measurement of protein cytokines in tissue extracts by enzyme-linked immunosorbent assays: application to lipopolysaccharide-induced differential milieu of cytokines.

OBJECTIVES: Determination of protein cytokines in local tissues would help to evaluate their local role in health, sickness behavior and immune-mediated diseases. Therefore, developing a simple quantitative method of protein cytokines in tissues/organs is highly important. METHODS: Mouse tissues were collected following intraperitoneal administration of endotoxin-free PBS or lipopolysaccharide. A mild detergent, 0.1% Igepal, was added in a buffer to enhance cytokines extraction. The tissues were then disrupted, homogenized, centrifuged and the supernatants were collected and assayed using solid-phase immunoassays. RESULTS: The presence of 0.1% Igepal extracted significantly more TNF-alpha from liver (322%: p<0.01), brain (358%: p<0.05), lungs (1600%: p<0.01), and more IL-10 from liver (220%: p<0.001), brain (4650%: p<0.001) than PBS alone. On the other hand, using 0.1% Igepal did not increase IFN-gamma extraction from liver, spleen, brain, lungs, skin and kidneys more than PBS alone. Furthermore, i.p. administration of LPS induced a differential milieu of cytokines. LPS increased significantly the production of TNF-alpha, IFN-gamma, and IL-10 from liver (521%, 123%, 72%: p<0.01, 0.04, 0.04), brain (470%, 122%, 280%: p< 0.01, 0.03, 0.01), peritoneal lavage (p<0.001) and blood (p<0.001). However, the pattern of increase was different for the above cytokines in spleen, skin, lungs and kidneys. CONCLUSIONS: The extraction of protein cytokines from tissues was superior with addition of mild detergent. Furthermore, our results showed a differential cytokines response to LPS with respect to tissue and cytokine type. This method should provide an important tool for studying local protein cytokines in behavioral pattern, sickness behavior, and immune-mediated diseases as well as to determine local therapeutic efficacy of immunomodulatory drugs.

The antimicrobial activities of Psidium guajava and Juglans regia leaf extracts to acne-developing organisms.

This study aims to present the in vitro inhibitory effect of Psidium guajava and Juglans regia leaf extracts on the main developer of acne lesions, Propionibacterium acnes (P. acnes), and other organisms that are isolated from acne lesions. Thirty-eight subjects (males and females) who had various types of acne were enrolled in the study. The contents of the acne lesions were cultured and the frequency of P. acnes (alone and with Staphylococci spp.) was 47%, whereas the frequencies for Staphylococcus aureus and Staphylococcus epidermidis were 13% and 24%, respectively. The antimicrobial activities of Psidium guajava and Juglans regia leaf extracts, determined by disk diffusion method (zone of inhibition), were compared to tea tree oil (TTO), doxycycline and clindamycin antibiotics. The zones of inhibition due to the Psidium guajava and Juglans regia leaf extracts ranged from 15.8-17.6 mm against P. acnes, 11.3-15.7 mm against S. aureus and 12.9-15.5 mm against S. epidermidis, respectively. These zones of inhibition were significantly higher than those of TTO and equivalent in case of Staphylococci spp., but less in case of P. acnes, to those obtained from doxycycline or clindamycin. It can be concluded that Psidium guajava and Juglans regia leaf extracts may be beneficial in treating acne especially when they are known to have anti-inflammatory activities.

Glucosamine enhances paracetamol bioavailability by reducing its metabolism.

Paracetamol has an extensive first-pass metabolism that highly affects its bioavailability (BA); thus, dose may be repeated several times a day in order to have longer efficacy. However, hepatotoxicity may arise because of paracetamol metabolism. Therefore, this project aimed to increase paracetamol BA in rats by glucosamine (GlcN). At GlcN-paracetamol racemic mixture ratio of 4:1 and paracetamol dose of 10 mg/kg, paracetamol area under the curve (AUC) and maximum concentration (Cmax ) were significantly increased by 99% and 66%, respectively (p < 0.05). Furthermore, paracetamol AUC and Cmax levels were increased by 165% and 88% in rats prefed with GlcN for 2 days (p < 0.001). Moreover, GlcN significantly reduced phase Ι and phase I/ΙΙ metabolic reactions in liver homogenate by 48% and 54%, respectively. Furthermore, GlcN molecule was found to possess a good in silico binding mode into the CYP2E1 active site-forming bidentate hydrogen bonding with the Thr303 side chain. Finally, serum ALT and AST levels of rats-administered high doses of paracetamol were significantly reduced when rats were prefed with GlcN (p < 0.01). In conclusion, GlcN can increase the relative BA of paracetamol through reducing its metabolism. This phenomenon is associated with reduction in hepatocytes injury following ingestion of high doses of paracetamol.

The effect of estradiol, but not progesterone, on the production of cytokines in stimulated whole blood, is concentration-dependent.

OBJECTIVES: The purpose of this study was to determine the effects of estradiol and progesterone on interferon-gamma (IFN-gamma), interleukin (IL)-12, IL-10 and tumor necrosis factor-alpha (TNF-alpha) productions in polyclonal activators (phytohemagglutinin+lipopolysaccharide)-stimulated whole blood cultures. METHODS: Nineteen healthy males and females volunteered in the study. Blood samples were drawn, diluted, and cultured for 24h with different concentrations of estradiol, progesterone or hydrocortisone and then PHA+LPS was added for another 24 h. The supernatant, then, was harvested and assayed for IL-12 p70, IFN-gamma, IL-10 and TNF-alpha. RESULTS: At preovulatory concentrations, estradiol enhanced significantly IFN-gamma, IL-12 and IL-10, but not TNF-alpha, production levels and reversed the suppressive effect of hydrocortisone in PHA+LPS stimulated whole blood. While IL-10 levels kept increasing at pregnancy estradiol concentrations, IFN-gamma, IL-12 levels and IFN-gamma/IL-10 ratio decreased significantly. No effect of progesterone on IL-12 p70, IFN-gamma, IL-10 and TNF production levels was observed. CONCLUSIONS: The present study shows that those pregnancy estradiol concentrations (and higher) enhance the production of IL-10 and reduce IL-12, IFN-gamma levels and IFN-gamma/IL-10 ratio in stimulated whole blood cells. Because of the known IL-10 inhibitory actions on T helper (Th) 1 cells and monocytes/macrophages, these high IL-10 levels keep Th2 cytokines favored during pregnancy and may be useful in shifting Th1-mediated autoimmune diseases towards non-pathogenic Th2 pathway.