Marc C. Karam

The antioxidant and anticancer effects of Wild Carrot oil extract

Daucus carota L. ssp. carota (Apiacea) is used in traditional medicine in Lebanon and in different regions throughout the world. The present study investigates the in vitro anticancer activities of Daucus carota oil extract (DCOE) on four human cancer cell lines as well as its in vitro antioxidant activity. DCOE was extracted from the dried umbels with 50:50 acetone‐methanol. The oil extract was analyzed by gas chromatography–mass spectrometry and screened for its antioxidant properties in vitro using 1,1‐diphenyl‐2‐picryl hydrazyl free radical scavenging assay (DPPH), ferrous ion chelating assay (FIC) and the ferric reducing antioxidant power assay (FRAP). The anticancer activity of the oil extract against human colon (HT‐29, Caco‐2) and breast (MCF‐7, MDA‐MB‐231) cancer cell lines was evaluated using the trypan blue exclusion method and the WST‐1 cell proliferation assay. DCOE exhibited antioxidant activity in all assays used. The FRAP value was 164 ± 5.5 µmol FeSO4/g, and the IC50 values for DPPH and FIC assays were 2.1 ± 0.03 mg/ml and 0.43 ± 0.02 mg/ml, respectively. Also, DCOE demonstrated a significant increase in cell death and decrease in cell proliferation. The effect of DCOE on the cell lines exhibited time and dose‐dependent responses. The present study established that DCOE possesses both antioxidant and promising anticancer activities. Copyright

In Leishmania major-induced inflammation, interleukin-13 reduces hyperalgesia, down-regulates IL-1β and up-regulates IL-6 in an IL-4 independent mechanism.

Infection with high dose Leishmania major induces a sustained hyperalgesia in BALB/c mice while low dose induces a short lived hyperalgesia both accompanied with the upregulation of IL-1β and IL-6. Although IL-13 was shown to reduce the high dose L. major hyperalgesia during the treatment period, this effect was accompanied by a significant decrease in the levels of IL-1β and a significant increase in the levels of IL-6 in the paws of mice even beyond this period. Those results suggest that IL-13 exerts those effects via the induction of another mediator, IL-4 being a potential candidate due to its known hypoalgesic effects in other models and to its close functional closeness to IL-13 especially at the level of receptors. In this study we correlated the pain thresholds and the levels of IL-1β, IL-6 and IL-4 with the period of IL-13 treatment and beyond it in mice infected with high and low dose of L. major. The results of both models show that IL-1β plays no direct role in provoking the observed hyperalgesia after stopping the treatment with IL-13 which is in contrary to IL-6 which might be a key player after the treatment period. Furthermore we demonstrate that there is no correlation between the levels of IL-4, hyperalgesia, the decreased IL-1β levels and the increased levels of IL-6 in the paws of IL-13 treated and L. major (high and low dose) infected BALB/c mice.

Montivipera bornmuelleri venom selectively exhibits high cytotoxic effects on keratinocytes cancer cell lines

CONTEXT The Viperidae family venom is a rich source of bioactive compounds such as many proteases, which cause tissue necrosis and affect mostly the vascular system. However, the venom exhibits therapeutic potentials and has contributed to the development of some medical drugs. Specifically, the Montivipera bornmuelleri venom has shown to exhibit antibacterial, pro-inflammatory and antifungal activities. OBJECTIVE This work evaluates the cytotoxic effect of the M. bornmuelleri venom on human-derived keratinocytes including the non-tumorigenic HaCaT, the benign A5 and the low-grade malignant II4 cells. MATERIALS AND METHODS The toxicity of different venom concentrations (0.9, 1.87, 3.75, 7.5, 15, 30 and 60μg/mL) and their effect on the viability of the cells lines were assessed using the Lactate Dehydrogenase (LDH) activity and the Trypan blue tests after 24h of incubation. RESULTS The venom was able to reduce the viability of all cell lines in a dose dependent manner with the HaCat cells being the least affected. For example, the 60μg/mL dose induced a more significant decrease the viability of A5 (44%) and II4 (21.33%) keratinocytes as compared to HaCaT cells (70.63%). Also, this venom showed a higher cytotoxic activity on the A5 (52.45%) and II4 (98.67%) cells as compared to HaCaT cells (30.14%) with an IC50 estimated at 10μg/mL on II4 and at 60μg/mL on benign A5. DISCUSSION AND CONCLUSION Those differential cytotoxic effects of the M. bornmuelleri venom pave the road for more advanced studies which might unravel the potential anticancer effects of this venom.

Antioxidant and hepatoprotective activities of the oil fractions from wild carrot (Daucus carota ssp. carota)

Abstract Context: Wild carrot, Daucus carota L. ssp. carota (Apiacae), is widely distributed throughout the world and has various uses in traditional medicine in Lebanon. Objective: The present study aimed to fractionate and analyze the chemical composition of the Daucus carota oil extract (DCOE) fractions and to evaluate their antioxidant and hepatoprotective properties in vitro and in vivo. Materials and methods: DCOE was chromatographed on silica gel column to produce four fractions: pentane (F1), 50:50 pentane:diethyl ether (F2), diethyl ether (F3), and 93:7 chloroform: methanol (F4). Qualitative and quantitative analyses of oil fractions were performed by GC-MS and HPLC techniques. The in vitro antioxidant properties were assessed using DPPH, FIC, and ferric-reducing antioxidant power (FRAP) assays. The hepatoprotective property was determined by examining the levels of serum markers (alanine transaminase (ALT) and aspartate transaminase (AST)) and hepatic antioxidant (superoxide dismutase (SOD), catalase (CAT), and glutathione-S-transferase (GST)) enzymes in CCl4-intoxicated mice pretreated with intraperitoenal 50, 100, or 200 mg/kg b.w. of the oil fractions for 5 d. Results: GCMS analysis of F2 revealed the presence of 2-himachalen-6-ol (61.4%) which is reported for the first time in Daucus carota species. F3 and F4 were rich in phenolics and flavonoids and demonstrated significant DPPH activity (IC50 = 0.29 and 0.38 mg/ml, respectively) and high FRAP values (225.11 and 437.59 µmol FeSO4/g, respectively). The sesquiterpene-rich fraction F1 had the highest FIC ability (IC50 = 0.28 mg/ml). Pretreatment with F1 and F4 reversed the CCl4-induced decrease in SOD, CAT, and GST levels and reduced significantly hepatic damage. Discussion and conclusion: The current results suggested that wild carrot oil fractions exhibited a unique chemical composition and possessed significant antioxidant activities as well as hepatoprotective effects against CCl4-induced hepatotoxicity.

Atenolol Reduces Leishmania major-Induced Hyperalgesia and TNF-α Without Affecting IL-1β or Keratinocyte Derived Chemokines (KC)

Infection with a high dose of the intracellular parasitic protozoan Leishmania major induces a sustained hyperalgesia in susceptible BALB/c mice accompanied by up-regulation of the pro-inflammatory cytokines IL-1β and IL-6. Interleukin-13 (IL-13) has been shown to reduce this hyperalgesia (despite increased levels of IL-6) and the levels of IL-1β during and after the treatment period. These findings favor the cytokine cascade leading to the production of sympathetic amines (involving TNF-α and KC) over prostaglandins (involving IL-lβ and IL-6) as the final mediators of hyperalgesia. The aim of this study was to investigate the effect of daily treatment with the β-blockers atenolol on L. major-induced inflammation in mice with respect to hyperalgesia as well as the levels of TNF-α and KC (the analog of IL-8 in mice). Our data demonstrates that atenolol is able to reduce the L. major induced sustained peripheral hyperalgesia, which does not seem to involve a direct role for neither IL-lβ nor KC. Moreover, our results show that TNF-α may play a pivotal and direct role in sensitizing the peripheral nerve endings (nociceptors) since its level was reduced during the period of atenolol treatment, which correlates well with the reduction of the observed peripheral, but not central, hyperalgesia. These findings contribute to a better understanding of the cytokine cascade leading to hyperalgesia and may lead to the development of new and more efficient medications for many types of pain.

The combination of CRP isoforms with oxLDL decreases TNF‑α and IL‑6 release by U937‑derived macrophages

C-reactive protein (CRP) and oxidized low density lipoprotein (oxLDL) serve major roles at both early and advanced stages of atherosclerosis. CRP exists in two isoforms, monomeric (m) and pentameric (p), that bring about pro- or anti-inflammatory effects in macrophages. In addition, CRP may form a complex with oxidized low-density lipoprotein (oxLDL) via phosphatidylcholine, thus decreasing its pro-inflammatory effects within macrophages. The aim of the present study was to investigate the single and the combined effects of mCRP, pCRP and oxLDL on U937-derived macrophages. In the current study, U937-derived macrophages were treated in vitro with different combinations of CRP isoforms with or without oxLDL. The levels of major inflammatory cytokines [interleukin (IL)-1β, IL-6, IL-8 and tumor necrosis factor (TNF)-α] along with the production of reactive oxygen species (ROS) were determined. TNF-α and IL-6 levels were significantly decreased (P<0.05) by the effect of mCRP and pCRP combined with oxLDL. No significant changes were observed in IL-1β, IL-8 or ROS levels.

Montivipera bornmuelleri venom has immunomodulatory effects mainly up-regulating pro-inflammatory cytokines in the spleens of mice

Graphical abstract

Immunomodulatory effect of natural and modified Citrus pectin on cytokine levels in the spleen of BALB/c mice

Pectin is present in the cell wall of different vegetables and fruits. Beside its importance in the plant cell wall, pectin has enticed great attention for its beneficial effects on human health. It was shown to decrease cholesterol levels, to possess anti-oxidative, anti-bacterial and anti-cancer activity. The immunomodulatory activity of pectin and its mechanism of action is recently being investigated. In this study, the differential immunomodulatory activities of both CP (citrus pectin) and MCP (modified citrus pectin) were investigated. Females BALB/c mice (20-25 g) were randomly divided into 7 groups and different concentrations of CP and MCP (0%, 1.5%, 3% and 5%) were added to their drinking water for 21 days. Then, the splenic level of IL-1β, IL-4, IL-10, IL-17, IFN-γ and TNF-α were evaluated using ELISA. Both CP and MCP exhibited immunomodulatory activities by increasing the levels of the pro-inflammatory cytokines IL-17, IFN-γ and TNF-α levels. This tendency seems to be regulated by the up-regulation of IL-4 levels but with no major effect on those of IL-10. Therefore, CP and especially MCP have potential immunomodulatory effects which might be highly beneficial in immunotherapy.Pectin is present in the cell wall of different vegetables and fruits. Beside its importance in the plant cell wall, pectin has enticed great attention for its beneficial effects on human health. It was shown to decrease cholesterol levels, to possess anti-oxidative, anti-bacterial and anti-cancer activity. The immunomodulatory activity of pectin and its mechanism of action is recently being investigated. In this study, the differential immunomodulatory activities of both CP (citrus pectin) and MCP (modified citrus pectin) were investigated. Females BALB/c mice (20-25 g) were randomly divided into 7 groups and different concentrations of CP and MCP (0%, 1.5%, 3% and 5%) were added to their drinking water for 21 days. Then, the splenic level of IL-1β, IL-4, IL-10, IL-17, IFN-γ and TNF-α were evaluated using ELISA. Both CP and MCP exhibited immunomodulatory activities by increasing the levels of the pro-inflammatory cytokines IL-17, IFN-γ and TNF-α levels. This tendency seems to be regulated by the up-regulation of IL-4 levels but with no major effect on those of IL-10. Therefore, CP and especially MCP have potential immunomodulatory effects which might be highly beneficial in immunotherapy.

Vipers of the Middle East: A Rich Source of Bioactive Molecules

Snake venom serves as a tool of defense against threat and helps in prey digestion. It consists of a mixture of enzymes, such as phospholipase A2, metalloproteases, and l-amino acid oxidase, and toxins, including neurotoxins and cytotoxins. Beside their toxicity, venom components possess many pharmacological effects and have been used to design drugs and as biomarkers of diseases. Viperidae is one family of venomous snakes that is found nearly worldwide. However, three main vipers exist in the Middle Eastern region: Montivipera bornmuelleri, Macrovipera lebetina, and Vipera (Daboia) palaestinae. The venoms of these vipers have been the subject of many studies and are considered as a promising source of bioactive molecules. In this review, we present an overview of these three vipers, with a special focus on their venom composition as well as their biological activities, and we discuss further frameworks for the exploration of each venom.