Phytochemical Screening and in vitro Anticancer Activity of Some Medicinal Plants Grown in the Kingdom of Saudi Arabia

SUMMARY: This study evaluated the phytochemical screening, antioxidant capacity, and in vitro anticancer activities of four plants namely, Gypsophila capillaris, Anabasis lachnantha, Haloxylon salicornicum, and Horwoodia dicksoniae which belong to four different families: Caryophyllaceae, Amaranthaceae, Chenopodiaceae, Brassicaceae, respectively. The total phenolics, anthocyanins, saponins, total antioxidant capacity (TAC), and DPPH assays were determined by spectrophotometer. In vitro anticancer activity was assessed using two human cancer cell lines; hepatocellular carcinoma (HepG-2) and breast adenocarcinoma (MCF-7) to estimate the inhibition concentration 50 % (IC50). The results showed that H. dicksoniae has the highest concentrations of phenolics and saponins, while H. salicornicum has the highest DPPH. The highest concentration of TAC was found in G. capillaries. Among the tested extracts, G. capillaries and H. salicornicum have the potential activity against MCF-7 and HepG-2 cell lines in vitro. The content of polyphenols in G. capillaries was profiled by high-performance liquid chromatography (HPLC). The highest concentration among the phenolic compounds was chlorogenic (60.8 µg/ml) while the highest concentration among the flavonoid compounds was hesperidin (1444.92 µg/ml). In summary, G. capillaries and H. salicornicum extracts have potent anticancer activity against HepG-2 and MCF-7 cell lines.

Este estudio evaluó la detección fitoquímica, la capacidad antioxidante y las actividades anticancerígenas in vitro de cuatro plantas, Gypsophila capillaris, Anabasis lachnantha, Haloxylon salicornicum y Horwoodia dicksoniae, que pertenecen a cuatro familias diferentes: Caryophyllaceae, Amaranthaceae, Chenopodiaceae y Brassicaceae, respectivamente. Los ensayos de fenólicos totales, antocianinas, saponinas, capacidad antioxidante total (TAC) y DPPH se determinaron mediante espectrofotómetro. La actividad anticancerígena in vitro se evaluó utilizando dos líneas celulares de cáncer humano; carcinoma hepatocelular (HepG-2) y adenocarcinoma de mama (MCF- 7) para estimar la concentración de inhibición del 50 % (IC50). Los resultados indicaron que H. dicksoniae tiene las concentraciones más altas de fenólicos y saponinas, mientras que H. salicornicum tiene el DPPH más alto. La mayor concentración de TAC se encontró en G. capillaries. Entre los extractos probados, G. capillaries y H. salicornicum tienen actividad potencial contra líneas celulares MCF-7 y HepG-2 in vitro. El contenido de polifenoles en G. capillaries se perfiló mediante cromatografía líquida de alta resolución (HPLC). La concentración más alta entre los compuestos fenólicos fue clorogénica (60,8 µg/ml), mientras que la concentración más alta entre los compuestos flavonoides fue la hesperidina (1444,92 µg/ml). En resumen, los extractos de Gypsophila capillaris y H. salicornicum tienen una potente actividad anticancerígena contra las líneas celulares HepG-2 y MCF-7.

Gingerol and/or sorafenib attenuates the DAB-induced HCC and hepatic portal vein dilatation via ATG4/CASP3 and COIIV/COX-2/NF-κB expression.

Ginger (Gin) has numerous therapeutic properties. One of Gin's most potent components is 6-gingerol, a naturally occurring phenol. This study aimed to investigate the therapeutic impact of gingerol and/or sorafenib on the ATG4/CASP3 and COIIV/COX-2/NF-B Expression as a potential therapy for DAB-induced HCC. Gin was administered to HCC mice induced by p-Dimethylaminoazobenzene (DAB) alone or combined with sorafenib (Sor). Superoxide dismutase (SOD), catalase (CAT), and oxidative stress malondialdehyde (MDA), as well as biochemical markers including AST, ALT, ALP, Albumin, and Bilirubin, were examined. The expression of oncogenes (COIIV, COX-2, NF-κB, and survivin) and tumor suppressor genes (ATG4 and CASP3) was evaluated using qPCR. According to the results, the levels of MDA have been markedly decreased, while SOD and CAT have been increased. Further, the expression levels of tumor suppressor genes were upregulated, whereas the expression levels of oncogene genes were downregulated. Furthermore, in a dose-dependent manner, gingerol has shown the potential to alleviate hepatic portal vein (PV) dilatation and could offer a reliable therapy for HCC. This suggests combining the two compounds may be more effective than alone and that Gin could be a promising therapeutic option for HCC. The binding of Gin and Sor to the active sites of the target genes prevents them from functioning normally, which in turn stops the pathways from carrying out their oncogenic functions. Additionally, COX-2 inhibition reduces the production of certain pro-inflammatory compounds, which further averts oncogenesis. Conclusively, this study indicated that Gin has cytoprotective properties and anti-cancer activity that may be related to controlling oxidative stress. This effect may be achieved by suppressing the COIIV/COX-2/NF-κB pathway and upregulating the ATG4 /CASP3 pathways.

Artemisia annua L. (Sweet wormwood) leaf extract attenuates high-fat diet-induced testicular dysfunctions and improves spermatogenesis in obese rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Artemisia annua L., known as "sweet wormwood," is widely used in Egyptian folk medicine. Egyptians implement the aerial parts in the treatment of respiratory, digestive and sexual dysfunctions. However, the mechanism by which Artemisia annua improves testicular function is still being discovered. AIM OF THE STUDY: This study aimed to evaluate the modulatory effects of the crude leaf extract of Artemisia annua (AAE) on a high-fat diet induced testicular dysfunction in rats and compare it with the antilipolytic drug Orlistat. MATERIAL AND METHODS: Forty adult rats were randomly classified and assigned to four groups. The first group typically consumed a balanced diet and served as a negative control (GP1). A high-fat diet-induced obesity was applied to the other three groups for 12 weeks. A positive control remained on HFD for another 8 weeks, which is GP2. Other groups were administered for 8 consecutive weeks either with Orlistat (50 mg/kg body weight) or AAE (100 mg/kg body weight), which have been defined as GP3 and GP4, respectively. Testosterone (TST), follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were determined in the sera of all groups. In addition, the oxidant/antioxidant biomarkers such as protein carbonyl, malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT) activities, lactate dehydrogenase (LDH) and creatine kinase isoenzyme-B (CK-MB) were determined. An immunohistochemical stain with the apoptotic marker caspase-3 and the proliferating cell nuclear antigen (PCNA) were also investigated. RESULTS: In the testes of the obese group, the results showed hormonal imbalance, an increase in oxidative stress biomarkers and apoptosis. In the group treated with orlistat (GP3), noticeably more perturbations were noted. The obese rats that had been treated with AAE (GP4) showed a significantly reduced level of oxidative stress, hormonal balance restoration and reduced apoptosis. CONCLUSIONS: The crude leaf extract of A. annua is a potential herbal therapeutic for the treatment of obesity-related testicular dysfunction and the restoration of hormonal imbalance in obese rats.

Modulation of the oxidative damage, inflammation, and apoptosis-related genes by dicinnamoyl-L-tartaric acid in liver cancer.

Cancer cells can become resistant to existing treatments over time, so it is important to develop new treatments that target different pathways to stay ahead of this resistance. Many cancer treatments have severe side effects that can be debilitating and even life-threatening. Developing drugs that can effectively treat cancer while minimizing the risks of these side effects is essential for improving the quality of life of cancer patients. The study was designed to explore whether the combination of dicinnamoyl-L-tartaric (CLT) and sorafenib ((SOR), an anti-cancer drug)) could be used to treat hepatocellular carcinoma (HCC) in the animal model and to assess whether this combination would lead to changes in certain biomarkers associated with the tumour. In this study, 120 male mice were divided into 8 groups of 15 mice each. A number of biochemical parameters were measured, including liver functions, oxidative stress (malondialdehyde, (MDA); nitric oxide (NO)), and antioxidative activity (superoxide dismutase (SOD), and glutathione peroxidase (GPx)). Furthermore, the hepatic expressions of Bax, Beclin1, TNF-α, IL1ß, and BCl-2 genes were evaluated by qRT-PCR. The combination of SOR and CLT was found to reduce the levels of liver enzymes, such as AST, ALT, ALP, and GGT, and reduce the pathological changes caused by DAB and PB. The upregulation of TNF-α, IL1ß, and Bcl-2 genes suggests that the CLT was able to initiate an inflammatory response to combat the tumor, while the downregulation of the Bax and Beclin1 genes indicates that the CLT was able to reduce the risk of apoptosis in the liver. Furthermore, the combination therapy led to increased expression of cytokines, resulting in an enhanced anti-tumor effect.

Diarylheptanoids/sorafenib as a potential anticancer combination against hepatocellular carcinoma: the p53/MMP9 axis of action.

Hepatocellular carcinoma (HCC) is a serious and potentially fatal form of cancer associated with liver damage. New anticancer drugs are increasingly needed due to the increasing number of cancer cases every year. In this study, diarylheptanoids (DAH) from Alpinia officinarum were examined for their antitumor activity against DAB-induced HCC in mice, as well as their ability to reduce liver damage. Assays for cytotoxicity were conducted using MTT. The DAB-induced HCC Swiss albino male mice were given DAH and sorafenib (SOR) either as single treatments or in combination, and the effects on tumour development and progression were monitored. Malondialdehyde (MDA) and total superoxide dismutase (T-SOD) were evaluated along with biomarkers of liver enzymes (AST, ALT, and GGT). The apoptosis-related gene (CASP8), the apoptosis-related gene (p53), the anti-inflammatory genes (IL-6), the migration-related gene matrix metalloprotease-9 (MMP9), and the angiogenesis-related gene vascular endothelial growth factor (VEGF) were assessed using qRT-PCR in the hepatic tissue. As a final step, DAH and SOR were docked with CASP8 and MMP9 via molecular docking to propose potential mechanisms of action. Our results revealed that the combination of DAH and SOR has a potent inhibitory effect on the growth and viability of the HepG2 cell line. The outcomes demonstrated that DAH and SOR-treated HCC-bearing mice displayed a reduction in the tumour burden and liver damage as demonstrated by (1) parameters of repaired liver function; (2) low levels of hepatic MDA; (3) elevated levels of hepatic T-SOD; (4) p53, IL-6, CASP8, MMP9, and VEGF downregulation; and (5) enhanced hepatic structure. The best results were revealed in mice that were co-treated with DAH (given orally) and SOR (given intraperitoneally). The docking study also proposed that both DAH and SOR could inhibit CASP8 and MMP9's oncogenic activities and had a high affinity for these enzymes. In conclusion, according to study findings, DAH enhances SOR antiproliferative and cytotoxic effects and identifies their molecular targets. Furthermore, the results revealed that DAH was able to boost the anticancer effects of the drug SOR and reduce liver damage caused by HCC in mice. This suggests that DAH could be a potential therapeutic agent against liver cancer.

Amygdalin potentiates the anti-cancer effect of Sorafenib on Ehrlich ascites carcinoma and ameliorates the associated liver damage.

The burden of cancer diseases is increasing every year, therefore, the demands to figure out novel drugs that can retain antitumor properties have been raised. This study aimed to investigate the anti-tumor properties of amygdalin (Amy) against Ehrlich ascites carcinoma (EAC) bearing mice and its protective properties against liver damage. Amy and the standard anticancer drug Sorafenib (Sor) were given alone or in combination to Swiss albino female mice that had been injected with EAC cells. Biochemical parameters of liver function (AST, ALT, GGT, total protein, albumin), tumor volume, oxidative stress [malondialdehyde, (MDA)] and antioxidative [superoxide dismutase (SOD), and reduced glutathione (GSH)] markers were measured. The hepatic expression of the antioxidant-related gene [nuclear factor erythroid-2-related factor 2 (Nrf2)], the migration-related gene [matrix metalloprotease 9 (MMP9)], and the angiogenesis-related gene [vascular endothelial growth factor (VEGF)] were evaluated by qPCR. The results revealed that EAC-bearing mice treated with Amy and/or Sor showed a decrease in the tumor burden and hepatic damage as evidenced by (1) decreased tumor volume, number of viable tumor cells; (2) increased number of dead tumor cells; (3) restored the liver function parameters; (4) reduced hepatic MDA levels; (5) enhanced hepatic GSH and SOD levels; (6) upregulated expression of Nrf2; (7) downregulated expression of MMP9 and VEGF, and (8) improved hepatic structure. Among all treatments, mice co-treated with Amy (orally) and Sor (intraperitoneally) showed the best effect. With these results, we concluded that the Amy improved the antitumor effect of Sor and had a protective role on liver damage induced by EAC in mice.

Musa sp. Leaves Extract Ameliorates the Hepato-Renal Toxicities Induced by Cadmium in Mice.

Heavy metals intoxication causes several health problems that necessitate finding new protective and therapeutic approaches. This study aimed to evaluate the impact of Musa sp. leaves extract (MLE) on hepato-renal toxicities induced by cadmium (Cd) in male mice. The phytochemical screening, metal chelating activity (MCA), and the median lethal dose (LD50) of MLE were determined. Fifty CD-1 male mice were used and intraperitoneally (i.p.) injected with MLE (1000 to 5000 mg/kg b.wt) for MLE LD50 determination. Another 50 mice were used for evaluating the effect of MLE on Cd toxicity. Blood samples were collected for hematological, liver, and kidney functions assessments. Liver tissue homogenates were used for determination of oxidant/antioxidant parameters. Liver and kidney tissues were harvested for histopathological and molecular investigations. MLE showed potent in vitro antioxidant activities. The MCA and LD50 of the MLE were 75 µg/mL and 3000 mg/kg b.wt, respectively. MLE showed beneficial therapeutic activity against hepato-renal toxicities in Cd-intoxicated mice, evidenced by improving the hematological, biochemical, histopathological, and molecular alterations.

Metformin Causes Hepato-renal Dysfunctions in Obese Male Rats

Abstract Obesity results in detrimental effects on different body organs. Metformin (Met) has described to decrease the body weight of obese patient and to control the glucose level. This study aimed to evaluate the role of Met treatment for long period on the functionality of liver and kidney organs of obese rats. Forty rats were used in this study and divided into four groups as the following: group 1 (Gp1) was served as a negative control that administered orally with 200 µL of H2O. Gp2 of rats was administered with Met (200 mg/kg) daily for a month. Gp3 was obese rats, and Gp4 was obese rats administered with Met as in Gp2. All rats were sacrificed to analyze hematological, biochemical, and histopathological changes. The results showed that Met decreased the body weight of both naïve and obese rats, however, it caused hepato-renal dysfunctions in obese rats as evidenced by increased the levels of ALT, AST, urea, creatinine, and MDA and decrease in the antioxidants biomarkers (SOD, Cat and GSH). Collectively, Met causes liver and kidneys dysfunctions of obese rats and is not recommended to described for obese persons.

Toxicity of bean cooking media containing EDTA in mice.

The possible renal and hepatic toxicities of ethylenediaminetetraacetic acid (EDTA) in bean cooking media were studied using 100 male albino mice. Two sublethal doses of EDTA were used to explore their toxic effects; 20 mg/kg and 200 mg/kg, which corresponded to 1/100th and 1/10th of LD50, respectively. Accordingly, the toxicity study was performed using 50 mice, divided into five groups (n = 10/group) as follows: group 1 (Gp1) served as a negative control and was orally administered normal saline; group 2 (Gp2) was administered the bean cooking medium; group 3 (Gp3) was administered EDTA (200 mg/kg); group 4 (Gp4) was administered bean cooking medium containing 20 mg/kg of EDTA; and group 5 (Gp5) was administered bean cooking medium containing 200 mg/kg of EDTA. The results showed no significant changes in liver and kidney functions in Gp2 while Gp3, Gp4, and Gp5 exhibited significant increases in adverse liver and kidney function markers. Hematocrit values were significantly decreased in Gp3 and Gp5, while the total white blood cells counts were significantly decreased in Gp3 and significantly increased in Gp5. The number of platelets was decreased in Gp3, Gp4, and Gp5. The blood levels of sodium (Na+), iron (Fe2+), and calcium (Ca2+) were decreased in Gp3, Gp4, and Gp5 due to the chelating effects of EDTA. The hepatic and renal architectures were disorganized in Gp3, Gp4, and Gp5 with some hemorrhagic manifestations in livers and kidneys of mice. These results demonstrate that EDTA in bean cooking is harmful in mice under the conditions of this study, and the potentially harmful effects in humans supports restricting its use.

The Toll-Like Receptor 3 Agonist Polyriboinosinic Polyribocytidylic Acid Increases the Numbers of NK Cells with Distinct Phenotype in the Liver of B6 Mice.

One of the activating factors of the cells of the innate immune system is the agonists of toll-like receptors (TLRs). Our earlier publications detailed how poly(I:C), a TLR3 agonist, elevates the NK cell population and the associated antigen-specific CD8+ T cell responses. This study involved a single treatment of the B6 mice with poly(I:C) intraperitoneally. To perform a detailed phenotypic analysis, mononuclear cells were prepared from each of the liver, peripheral blood, and spleen. These cells were then examined for their NK cell population by flow cytometric analysis following cell staining with indicated antibodies. The findings of the study showed that the NK cell population of the liver with an NK1.1highCD11bhighCD11chigh B220+Ly6G- phenotype was elevated following the treatment with poly(I:C). In the absence of CD11b molecule (CR3-/- mice), poly(I:C) can still increase the remained numbers of NK cells with NK1.1+CD11b- and NK1.1+Ly6G- phenotypes in the liver while their numbers in the blood decrease. After the treatment with anti-AGM1 Ab, which induced depletion of NK1.1+CD11b+ cells and partial depletion of CD3+NK1.1+ and NK1.1+CD11b- cell populations, poly(I:C) normalized the partial decreases in the numbers of NK cells concomitant with increased numbers of NK1.1-CD11b+ cell population in both liver and blood. Regarding mice with a TLR3-/- phenotype, their injection with poly(I:C) resulted in the partial elevation in the NK cell population as compared to wild-type B6 mice. To summarise, the TLR3 agonist poly(I:C) results in the elevation of a subset of liver NK cells expressing the two myeloid markers CD11c and CD11b. The effect of poly(I:C) on NK cells is partially dependent on TLR3 and independent of the presence of CD11b.

Antitumor efficacy of EDTA co-treatment with cisplatin in tumor-bearing mice

Ethylenediamine tetraacetic acid (EDTA) is used in various medical applications. The aim of this study is to investigate the antitumor efficacy of EDTA alone or with cisplatin (Cis). Fifty male albino mice were used to assess the median lethal dose (LD50) of EDTA via intraperitoneal (i.p) injection. To determine the antitumor activity, fifty female albino mice were divided into five groups as the following; Group 1 (Gp1) was negative control; (Gp2-5) inoculated i.p with 2×106 Ehrlich Ascites Carcinoma (EAC) cells/mouse. After one day, Gp3, Gp4 and Gp5 injected with Cis (2 mg/kg), EDTA (25 mg/kg) and Cis (2 mg/kg)/EDTA (25 mg/kg) for six days, respectively. At day 14, all groups were sacrificed to assess the tumor profile, liver enzymes (alanine transaminases and aspartate transaminases), kidney function (urea and creatinine) and electrolytes (Na+, K+ and Ca2+). The results showed that the i.p LD50 of EDTA was 250 mg/kg. Treatment with EDTA alone did not show any antitumor activity and did not interfere with the antitumor efficacy of Cis. Biochemical findings revealed that EDTA had mild toxicity on liver and kidneys functions. In summary, EDTA had no antitumor effect and did not alter the Cis efficacy.

Cooking with EDTA reduces nutritional value of Vicia faba beans.

Ethylenediamine terta-acetic acid (EDTA) used to accelerate the cooking process of Vicia (V. faba) beans. In this study, the effect of cooking with EDTA on the nutritional value of V. faba beans was addressed. Water contents, total proteins, lipids, carbohydrates, minerals and amino acids were determined before and after boiling with EDTA (2 g/L). In both of whole beans and seed coats, the water content was increased after boiling with EDTA. In contrast, the levels of proteins, lipids and carbohydrates were significantly decreased in both the whole beans and seed coats upon boiling with EDTA. Furthermore, the levels of sodium were increased while, the levels of other minerals were decreased. All amino acids were significantly decreased in the whole beans and increased in the seed coats after boiling with EDTA. EDTA addition to V. faba beans during the cooking process decreased the nutritional value of the cooked V. faba beans.

Cardioprotective effect of green tea extract and vitamin E on Cisplatin-induced cardiotoxicity in mice: Toxicological, histological and immunohistochemical studies.

BACKGROUND: Cisplatin (CP) has been used in wide range for cancer treatment. Although nephrotoxicity of CP was the main complication, cardiotoxicity has been reported. OBJECTIVES: This study investigates the protective role of green tea extract (GTE) and vitamin E (Vit-E) against CP-induced cardiotoxicity, and assesses their impact on CP antitumor efficacy. MATERIALS AND METHODS: Forty-eight male albino Balb/c mice were randomly divided into six groups, 8 per/group (Gp) were included. Gp1 served as control; Gp2 and Gp3 received oral GTE (400 mg/kg) and Vit-E (100 mg/kg) for 30 consecutive days respectively. Gp4 had received CP (7 mg/kg i.p.) once on the 27th day; Gp5 had received GTE (400 mg/kg p.o.) for 30 days and CP (7 mg/kg i.p.) on the 27th day. Gp6 had received Vit-E (100 mg/kg p.o.) for 30 days and CP (7 mg/kg i.p.) on the 27th day. Blood and tissues samples were harvested for biochemical and histopathological investigations. To evaluate the effect of GTE and Vit-E on the antitumor efficacy of CP, 49 female albino mice were inoculated intraperitoneally by Ehrlich ascetic carcinoma -cells (2 × 106/mouse) then treated with none, corn oil, CP, CP/GTE, CP/Vit-E, GTE or Vit-E. RESULTS: CP injection significantly increased Troponin I, CPK, CK-MB, malondialdehyde (MDA), and nitric oxide (NO) levels, while glutathione peroxidase (GPx), superoxide dismutase (SOD) and catalase levels were significantly reduced with disruption of cardiac muscle fibers, loss of striations, absence of intercalated disc, and the nuclei are pyknotic. Treatment with GTE and Vit-E improve the biochemical and histological parameters. Treatment with CP alone led to eradication of the tumor cells from the tumor-bearing mice. However, co-administration of GTE or Vit-E orally with CP did not interfere with its therapeutic effects. CONCLUSION: Treatment with GTE and Vit-E significantly ameliorated the CP cardiotoxicity and improved the myocardial histopathological architecture. GTE and Vit-E may be combined with CP to alleviate cardiotoxicity in cancer chemotherapy without interfering with its antitumor activity.

Evaluation of antibacterial and anticancer properties of poly(3-hydroxybutyrate) functionalized with different amino compounds.

The chemical modification of biodegradable poly(3-hydroxybutyrate) (PHB) is useful for biomedical applications. In this study, the transesterification reaction of PHB was carried out under reflux conditions in the presence of 1,4-butanediol to form telechelic PHB-diol. Further modification of PHB-diol into PHB-diacrylate was carried out by the reaction of PHB-diol with acryloyl chloride. PHB-diacrylate was grafted with amino compounds such as 1,4-butanediamine, 1,3-propanediamine, 1,2-ethylenediamine, piperazine, cyclohexylamine, 2,2'-(ethane-1,2-diylbis(oxy)) diethanamine (jeffamine EDR 148) and morpholine via Michael-type addition reaction. The functionalized amino-PHB polymers were characterized by using FTIR and 1H NMR techniques. XRD showed that amino-PHB polymers have different crystallinity compared with neat PHB. Some biological activities of amino-PHB polymers were determined such as antibacterial, antioxidant and anticancer activities. In this regard, the results showed that PHB-ethylenediamine revealed a potent antibacterial activity against Staphlococcus aureus, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa. All amino-PHB polymers under the study showed reasonable antioxidant activity. Among these polymers, PHB-piperazine showed a potent anticancer effect against in vivo Ehrlich ascetic carcinoma bearing mice.

Enhancing Antitumor Efficacy of Cisplatin Low Dose by EDTA in Ehrlich Ascetic Carcinoma Bearing Mice

Abstract In a recent study, the treatment of different human cancer cell lines in vitro with ethylene diamine tetra-acetic acid (EDTA) showed a promising anticancer activity which could be a novel promising approach for cancer treatment. The aim of this study is to address the ability of EDTA to enhance the antitumor efficacy of the low dose of cisplatin (Cis) treatment in Ehrlich ascetic carcinoma (EAC) bearing mice. Sixty female albino mice were divided into six groups. The 1st group of mice was served as a negative control. 2nd - 6th groups were inoculated intraperitoneal (i.p) with 2×106 EAC cells/mouse. After one day of inoculation, the 2nd, 3rd and 4th groups were injected daily for 6 days (early treatment) with phosphate buffer saline, low dose of Cis and Cis/EDTA, respectively. After six days, the 5th and 6th groups were injected with the low dose of Cis and Cis/EDTA for 6 consecutive days (late treatment), respectively. At day 14, all groups of mice were sacrificed, sera were collected for biochemical assessment, then tumor volumes, counts, live and dead cells were determined from all groups. The results showed that EDTA co-treatment enhanced the efficacy of low dose of Cis at early and late time points. In addition, EDTA co-treatment potentially ameliorated the Cis-induced side effects on liver and kidney functions. In summary, co-therapy with EDTA could enhance the chemotherapeutic efficacy of low dose of Cis.

Cyclophosphamide eradicates murine immunogenic tumor coding for a non-self-antigen and induces antitumor immunity.

Although the majority of cancers respond to chemotherapy, most cancer types relapse, at least in part, due to the poor immunogenicity of most tumor. We have reported before that treatment of tumor bearing mice with a combination of the anti-cancer chemotherapy cyclophosphamide (CTX) and immunotherapy can result in complete tumor regression using T-cell receptor (TCR) transgenic CD8+ T cells specific to antigens. This study aimed to determine whether chemotherapy can cure immunogenic tumor which expresses non-self-tumor antigen and result in antitumor immunity. Either EL4 cell line, a poorly immunogenic thymoma, or EG7, a clone of EL4 cells transfected with ovalbumin (OVA), as a non-self-antigen were inoculated subcutaneously into wild type or splenectomized C57BL/6 mice and then treated once with intraperitoneal (i.p.) injection of 4 mg CTX/mouse. In certain experiments, the mice were rechallenged with the same tumor type 1-2 months after the primary challenge. Treatment of EL4 bearing mice with CTX induced transient antitumor effect followed by tumor progression. Interestingly, however, treatment of EG7-bearing mice with CTX resulted in regression of early and advanced tumors. EG7 tumor-free mice rejected the second and the third challenges with EG7 cells, but not with challenge EL4 cells. These antitumor effects did not require spleen, since splenectomized mice showed similar antitumor effects of CTX on EG7 cells. Taken together, these data indicate that expression of non-self-antigen by poorly immunogenic tumor might be a reliable means to increase its immunogenicity and its response to chemotherapy.

Growth, hydrolases and ultrastructure of Fusarium oxysporum as affected by phenolic rich extracts from several xerophytic plants.

Fusarium oxysporum, the causal agent of rot and wilt diseases, is one of the most detrimental phytopathogens for the productivity of many economic crops. The present study was conducted to evaluate the potentiality of some xerophytic plants as eco-friendly approach for management of F. oxysporum. Phenolic rich extracts from five plants namely: Horwoodia dicksoniae, Citrullus colocynthis, Gypsophila capillaris, Pulicaria incisa and Rhanterium epapposum were examined in vitro. The different extracts showed high variability in their phenolic and flavonoid contents as well as total antioxidant capacity. A strong positive correlation existed between the antifungal activity of the tested extracts and their contents of both total phenolics and flavonoids (r values are 0.91 and 0.82, respectively). Extract of P. incisa was the most effective in reducing the mycelial growth (IC50=0.92mg/ml) and inhibiting the activities of CMCase, pectinase, amylase and protease by 36, 42, 58 and 55%, respectively. The high performance liquid chromatography analysis of P. incisa extract revealed the presence of eight phenolic acids along with five polyphenolic compounds. The flavonol, quercetin and its glycosides rutin and quercetrin were the most abundant followed by the phenolic acids, t-cinnamic, caffeic, ferulic and vanillic. P. incisa extract not only affects the growth and hydrolases of F. oxysporum but also induces ultrastructure changes in the mycelium, as revealed by transmission electron microscopy. To our knowledge, this is the first study to investigate the mechanisms underlying the antifungal activity of P. incisa.

Higher Activities of Hepatic Versus Splenic CD8+ T Cells in Responses to Adoptive T Cell Therapy and Vaccination of B6 Mice with MHC Class-1 Binding Antigen.

The liver has unique microenvironment which is known to induce tolerance of cytolytic CD8+ T cells to hepatic and extra hepatic antigens, resulting in persistence of infection of the liver by the hepatitis B and C viruses. However, under some conditions, functional immune responses can be elicited in the liver in particular to show preferential retention of activated CD8+ T cells. It is not clear whether this retention depends on the type of the exogenous immunostimulatory or the endogenous innate immune cells. The T cell receptor (TCR) transgenic OT-1 (CD8+) mouse model was used in which OT-1 cells were harvested from the spleen of the donor and transferred into recipient mice followed by immunization with OVA peptide followed by injection of GM-CSF, CCL21 chemokine, or cytokines (IL-2, IL-12, or IL-15), or the toll-like receptor 3 agonist poly(I:C). Co-administration of any of these immunostimulatory agents relatively augmented the retention of CD8+ T cells with different levels of effects. Compared to spleen, the Ag-specific CD8+ T cells in the liver showed higher activities including expansion, proliferation, apoptosis and memory responses as well as cytolytic function. While depletion of natural killer cells significantly decreased the hepatic retention of the antigen-specific T cells, depletion of Kupffer cells showed opposite effect. Taken together, the antigen reactive T cells in the liver have higher activities than their counterparts in the peripheral tissues such as spleen. These data have important clinical implications for designing immunotherapeutic protocols toward the liver diseases.

Assessment of the Toxicity of Sub-chronic Low and High Doses of the Bio-insecticide Spinosad on the Liver, Kidney and the Cerebellum in Male Albino Mice

ABSTRACT Spinosad (SPD) is a highly selective insect control product. However, it was reported that SPD has toxicity toward other non-target organisms. This study was conducted to address the toxic effect of two sub-chronic low and high doses; 35 and 350 mg/kg SPD on some biochemical, histological and immunohistochemical parameters of the liver, kidney and cerebellum. Thirty-six male Swiss mice were divided into three groups of 12 mice each; first group (G1) served as a control, second group (G2) received a low sub-chronic dose of SPD that is equal to 35 mg/kg, and third group (G3) received a high sub-chronic dose of SPD that is equal to 350 mg/kg. The results showed that mice which were received 350 mg/kg SPD showed a significant decrease in the body weight and a significant increase in their relative kidney and spleen weights. They also showed a significant increase in alanine aminotransferase (ALT), triglycerides and urea levels. Histopathological examination showed cytoplasmic degeneration and cell necrosis in the liver and kidney. Immunohistochemical examination showed that cerebellum illustrated several neurodegenerative changes and a down-regulation of synaptophysin-Syp. In conclusion, exposure to a high dose of SPD that is equal to 350 mg/kg could cause a marked toxicity on the liver, kidney and cerebellum in male albino mice.