Hala E. Abdel-Hamied

Grape seed proanthocyanidin extract has potent anti-arthritic effects on collagen-induced arthritis by modifying the T cell balance.

Rheumatoid arthritis (RA) is an autoimmune disease characterised by chronic inflammation of the synovial joints, joint malformations, and disability. The continuous use of conventional anti-inflammatory drugs is associated with severe adverse effects. Grape seed proanthocyanidin extract (GSPE) is considered to have protective effects against several diseases. In this study based on the mouse adjuvant-induced-arthritis (AIA) model, we examined the effects of GSPE on the key mediators of arthritic inflammation, namely T cell subsets, glucocorticoid-induced tumour necrosis factor receptor (GITR) expressing cells, CD4(+)CD25(+)Foxp3(+) regulatory T (Treg) cells, Th17 cells, Th1/Th2 cytokines, and inflammatory mediator gene expression. We treated BALB/c mice with 25, 50, or 100 mg/kg GSPE or saline daily (14 days) per orally (p.o.) at the onset of AIA. At the peak phase of AIA (day 14), the heparinised whole blood and ankle joints of all groups were collected and tested. GSPE-treated mice showed a substantial reduction in the levels of T cell subsets, GITR-expressing cells, and Th1 cytokines as well as the inflammatory mediators (MCP-1, MIP-2, and ICAM-1) that induce them compared with the vehicle-treated (saline) and arthritic mice. However, GSPE significantly upregulated the number of Tregs and Th2 cytokine producing cell number or it also induced Th17/Treg rebalance and orchestrated various pro-inflammatory and anti-inflammatory cytokines and the gene expression of their mediators that mediate cellular infiltration into the joints. This might, contribute to its anti-arthritic activity. Our results suggest that p.o. treatment with GSPE attenuated AIA in mice might offer a promising alternative/adjunct treatment for RA.

Amelioration of autoimmune arthritis by naringin through modulation of T regulatory cells and Th1/Th2 cytokines.

Naringin, a well-known flavanone glycoside found in grapefruit and other citrus fruits, was determined to be an effective anti-inflammatory compound. We investigated the effect of naringin on the key mediators of arthritic inflammation, namely T cell subsets, CD4(+)GITR(+) expressing cells, CD4(+)CD25(+)Foxp3(+) (Treg), Th1/Th2 cytokines and inflammatory mediators. We treated Balb/c mice (p.o.) with naringin (20, 40 and 80 mg/kg) for 14 days. Compared with the vehicle-treated and arthritic-control mice, the naringin treatment demonstrated a considerable decrease in the level of T cells, CD4(+)GITR(+), Th1 cytokine and inflammatory mediator expressions. In contrast, naringin treatment resulted in significantly up-regulated Treg and Th2 cytokine levels. Therefore, the naringin-induced inhibition of the T cells, various pro-inflammatory cytokines and inflammatory mediators that facilitate cellular infiltration into the joints might have contributed to its anti-arthritic activity. Our data suggest that naringin diminished the AIA in mice and it could be a potential alternative/adjunct treatment for RA.

Thymoquinone suppression of the human hepatocellular carcinoma cell growth involves inhibition of IL-8 expression, elevated levels of TRAIL receptors, oxidative stress and apoptosis.

Abstract Hepatocellular carcinoma (HCC) is the fourth most common solid tumor worldwide. The chemokine interleukin-8 (IL-8) is overexpressed in HCC and is a potential target for therapy. Although the transcription factor NF-κB regulates IL-8 expression, and while thymoquinone (TQ; the most bioactive constituent of black seed oil) inhibits NF-κB activity, the precise mechanisms by which TQ regulates IL-8 and cancer cell growth remain to be clarified. Here, we report that TQ inhibited growth of HCC cells in a dose- and time-dependent manner, caused G2M cell cycle arrest, and stimulated apoptosis. Apoptosis was substantiated by activation of caspase-3 and -9, as well as cleavage of poly(ADP-ribose)polymerase. TQ treatments inhibited expression of NF-κB and suppressed IL-8 and its receptors. TQ treatments caused increased levels of reactive oxygen species (ROS) and mRNAs of oxidative stress-related genes, NQO1 and HO-1. Pretreatment of HepG2 cells with N-acetylcysteine, a scavenger of ROS, prevented TQ-induced cell death. TQ treatment stimulated mRNA expression of pro-apoptotic Bcl-xS and TRAIL death receptors, and inhibited expression of the anti-apoptotic gene Bcl-2. TQ enhanced TRAIL-induced death of HepG2 cells, in part by up-regulating TRAIL death receptors, inhibiting NF-κB and IL-8 and stimulating apoptosis. Altogether, these findings provide insights into the pleiotropic molecular mechanisms of TQ-dependent suppression of HCC cell growth and underscore potential of this compound as anti-HCC drug.

Role of a histamine 4 receptor as an anti-inflammatory target in carrageenan-induced pleurisy in mice

The histamine 4 receptor (H4R) is expressed primarily on cells involved in inflammation and immune responses. Despite much research into inflammatory diseases, no drugs with favourable safety profiles are yet available for their treatment. The aim of the present study was to determine the potential anti‐inflammatory effect of 4‐methylhistamine (4‐MeH) or JNJ77777120 (JNJ) and to explore the role of H4R in a mouse model of carrageenan (Cg) ‐induced pleurisy. A single dose of 4‐MeH or JNJ (30 mg/kg) was administered intraperitoneally 1 hr before Cg administration. The results illustrate that both the numbers of CD4+, CD25+, CD4+ CD25+, GITR+, GITR+ IL‐17A+‐expressing T cells and the levels of T helper type 1 (Th1)/Th17 cytokines were markedly increased in both the Cg‐treated and 4‐MeH‐treated groups, whereas the cytokines produced by Th2 cells were significantly decreased in the same groups. However, JNJ treatment significantly decreased both the number of T‐cell subsets and GITR+, GITR+ IL‐17A+‐expressing T cells, and the production of Th1/Th17 cytokines. Further, JNJ up‐regulated the expression of the Th2 cytokines. RT‐PCR analysis revealed an increased expression of interleukin‐1β, tumour necrosis factor‐α, monocyte chemoattractant protein‐1 and intercellular adhesion molecule‐1 in the Cg‐treated and 4‐MeH‐treated groups, which was reduced by treatment with JNJ in lung tissues. Moreover, histological examinations revealed anti‐inflammatory effects of JNJ, whereas 4‐MeH worsened Cg‐induced inflammation. In conclusion, the results of the present work clearly indicate that JNJ possesses important anti‐inflammatory properties that are increased in 4‐MeH‐treated mice, suggesting that H4R are involved in pleurisy and that JNJ has an anti‐inflammatory effect in associated disease conditions.

Involvement of histamine 4 receptor in the pathogenesis and progression of rheumatoid arthritis

Rheumatoid arthritis (RA) is one of the major autoimmune diseases with a global prevalence. Despite significant research into this disease, no drugs with acceptable safety profiles are yet available for its treatment. We investigated the possible anti-arthritic effects of the 4-methylhistamine (4-MeH) histamine 4 receptor (H4R) agonist and the JNJ77777120 (JNJ) H4R antagonist to explore the role of H4R in a mouse model of collagen antibody-induced arthritis (CAIA). Arthritis was induced via intravenous (tail vein) injection of Balb/c mice with a 5-clone cocktail of mAbs against collagen type II, followed by LPS, and the effects of treatment with 4-MeH or JNJ (30 mg kg(-1), i.p, twice daily) for 7 days (prophylactic or therapeutic regimens) were assessed. The results revealed increased paw edema, arthritic scores, joint histological inflammatory damage and matrix metalloproteinase-3 levels and high levels of Th1 pro-inflammatory cytokine mRNA and serum proteins in CAIA mice or following H4R activation via 4-MeH. Additionally, 4-MeH efficiently increased expression levels of NF-κB p65. JNJ-treated mice showed a substantial reduction in all the previously mentioned effects, with a similar trend being observed under prophylactic and therapeutic treatment regimens. The results of the present work indicate that JNJ exhibits significant anti-inflammatory and anti-arthritic activities, demonstrating the clear involvement of H4R antagonism in the pathogenesis and progression of RA.

Alpha-lipoic acid rebalances redox and immune-testicular milieu in septic rats

In the present study, lipopolysaccharide (LPS), as an immune modulator in male adult rats and alpha-lipoic acid (ALA), as a powerful biological antioxidant and anti-inflammatory, are examined to help understanding the role of the immune and redox perturbation in testicular dysfunction with a possible protection. A total of 60 male Swiss albino rats were divided into 5 groups (10/group) respectively as follows Saline, ALA-vehicle, ALA (200mg/kg), LPS (5mg/kg) started with 20 rats and LPS+ALA. Obtained data from previously reported study, in our laboratory, and from the present one revealed that LPS induced marked reductions in sperms count, motility and resulted in deterioration of the testicular histological features. In addition, LPS decreased testicular reduced glutathione (GSH) level and lactate dehydrogenase isoenzyme-x (LDH-x) activity. However, it increased testicular levels of malondialdehyde (MDA), nitric oxide (NO) and 8-hydroxydeoxyguanosine (8-HDG) in testicular DNA, along with increased serum IL-2 level. In contrast, rats pretreated with ALA showed almost complete normalization of all the tested parameters. In conclusion, LPS induced perturbation of the immune-testicular barrier as a result of redox imbalance with a subsequent testicular dysfunction. Pretreatment with ALA ameliorated all these effects by its immune-modulator and antioxidant mechanisms suggesting a protective role against male infertility in septic or severely infected patients.

Abstract 2528: Vardenafil is superior to idarubicin in treatment of glioma

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA Idarubicin (IDA) is a very potent second generation anthracycline that has shown high efficacy against glioma in vitro. However, this agent has failed to exert satisfactory therapeutic response in patients with brain tumors. Phosphodiesterase (PDE-5), an enzyme which hydrolyzes intracellular cyclic guanosine monophosphate (cGMP), is highly expressed in many brain tumor cell lines. We proposed that increasing cGMP in the brain by the potent PDE-5 inhibitor vardenafil (VAR) will augment the antitumor efficacy of IDA against glioma. We also proposed that VAR will induce apoptosis of glioma cells. Here we show for the first time that VAR (10 mg/kg) enhanced the antitumor activity of IDA (0.1mg/kg). VAR alone inhibited tumor growth and prolonged survival of glioma bearing rats. Histopathological examination revealed that tumors from glioma bearing rats treated with VAR alone or VAR+ IDA presented a significant reduction in mitotic index, indicating less proliferative tumors, as compared to those which received PBS (control) or IDA. Real time PCR results demonstrated that VAR inhibited mRNA expression of PDE-5 in the brains of animals treated with VAR, compared to control-treated group, implicating an important role for the inhibition of PDE-5 in the antitumor activity of VAR. Interestingly, the antitumor activity of VAR was similar to that of IDA. Moreover, animals receiving either VAR alone or VAR+IDA demonstrated statistically significant improvement in overall survival when compared with control (PBS) or IDA groups. In order to explain such an unexpected finding, we examined the antiproliferative, pro-apoptotic and anti-invasive activities VAR in vitro. MTT assay results showed that VAR significantly inhibited the growth of rat C6 glioma cells. Annexin V propidium iodide assay results revealed that the inhibition of C6 cell growth is mediated at least in part by inducing C6 apoptosis. Wound healing and soft agar colony formation assays showed that VAR inhibited the migration and anchorage-independent growth of C6 cells, respectively. This suggests that the enhanced antitumor effect of VAR could be due to a combined antiproliferative, pro-apoptotic and anti-invasive activities. Together, our data support anti-glioma properties of VAR, and provide a proof-of-concept basis for further development of VAR as potential adjuvant for treatment of brain tumors. Citation Format: Abdelkader E. Ashour, Abdulrahman Z. Alzahrani, Hala E. Abdel-Hamied, Khairy M. Zoheir, Sheikh F. Ahmed, Salem S. Al-Rejaie, Adel R. Abd-Allah. Vardenafil is superior to idarubicin in treatment of glioma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2528. doi:10.1158/1538-7445.AM2015-2528