Pistacia lentiscus L. revealed in vitro anti-proliferative activity on MCF-7 breast cancer cells and in vivo anti-mammary cancer effect on C57BL/6 mice through necrosis, anti-inflammatory and antioxidant enhancements

Abstract

Inflammation and oxidative stress are two interconnected processes that play a role in cancer development and progression. In the present research, we aimed to evaluate the anticancer effect of Pistacia lentiscus L. (PL) essential oil (EO) in vitro against MCF-7 breast cancer cells and in vivo in DMBA-mammary cancer induction on female C57BL/6 mice model as well as to investigate its anti-inflammatory and antioxidant potential as implicated mechanism. Our results revealed a new chemotypes-profile of 39 bio-compounds of PL EO. The main chemotypes were terpenoid and ketone compounds. In vitro, PL EO had a potent anti-proliferative activity against MCF-7 cells. In vivo, PL reduced the tumor number, volume, weight and burden values as compared to the DMBA-positive control group (p<0.05). Histopathology data confirmed the protective effect of PL traduced by the presence of necrosis area. PL EO revealed improvement on inflammatory perturbation in the C-RP levels and the complete blood cell count. Finally, PL improved oxidative disorders of lipid peroxidation, thiol groups, hydrogen peroxide and antioxidant enzymes depletion in plasma and mammary tissues. Also, a potent plasma scavenging capacity has been detected. Our data suggested that PL chemotypes inhibited cell proliferation, exerting a potential protective effect against DMBA-mammary cancer through anti-inflammatory and antioxidant enhancements. Targeting inflammation and oxidative stress may represent a promising strategy for breast cancer prevention and treatment.

Fig 1

Effect of essential oil of Pistacia lentiscus on MCF-7 cell viability (1A) and cell proliferation (1B). Cells were treated with increasing doses of PL EO extract for 24h and 72h. Cell viability was estimated by metabolic rate using the MTT assay. Absorbance values were measured at 560 nm wavelength and compared to untreated cells (containing medium alone). Data represent the mean ± standard error of mean (SEM) of two independent experiments performed in triplicates.

Fig 2. Pistacia lentiscus essential oil protective effect on DMBA-mammary cancer in C57Bl/6 female mice.

Twenty four C57Bl/6-female mice were treated as described in the experimental design of Section Material and Method: All animals were pre-treated with two doses of DMBA (20 and 50mg/kg, b.w. dose) associated with progesterone and estrogens treatments until mammary tumor induction. GroupI: had not received any treatment after DMBA-mammary cancer induction, while group II and III received PL EO (280 and 570 mg kg, b.w. dose. respectively) after DMBA mammary cancer induction. (2A). Measurement of body weight gain (2B) and the volume of mammary tumours growth (2C) were measured during the whole process for 5 months as described in Measurement of body weight, tumor volume and burden of Section Material and method. The tumours’ weight per mouse in each group (2D) was measured after euthanasia. The average numbers of mammary tumours in each mouse and the mean of tumour burden (2E) were calculated in various tumour diameter groups at the end of the process. Values are given as median and (minimum value- maximum value), *p<0.001: Compare to group I, positive control DMBA (ANOVA test) and # p<0.001: Compare to group II (ANOVA test).

Fig 3. Protective effect of Pistacia lentiscus essential oil on carcinogenesis process induced mammary cancer in female C57Bl/6 mice.

Microscopic observation (Real picture on the left, X10 on the middle, X40 on the right) in the mammary tissues of group I: DMBA positive control, Group II: DMBA then PL-1(280 mg/kg-b.w. dose) (CTR⁺ +PL-1) and Group III: DMBA then PL-2 (560 mg/kg-b.w. dose) (CTR⁺ + PL-2). All animals were pre-treated with two doses of DMBA (20 and 50mg/kg, b.w. dose) associated with progesterone and estrogens treatments until mammary tumor induction. GroupI: had not received any treatment after DMBA-mammary cancer induction, while group II and III received PL EO (280 and 570 mg/ kg, b.w. dose. respectively) after DMBA mammary cancer induction. Red circle and black arrow: necrosis areas; yellow arrow: atypical and pleomorphic mammary cells.

Fig 4. Effect of Pistacia lentiscus essential oil on 7,12-dimethylbenz(a)anthracene, inducing oxidative disorders in the mammary tissue and plasmaof female C57Bl/6 mice.

All animals were pretreated with two doses of DMBA (20 and 50mg/kg, b.w. dose) associated with progesterone and estrogens treatments until mammary tumor induction. GroupI: had not received any treatment, while group II and III received PL EO (280 and 570 mg/kg, b.w. dose, respectively). Results are represented by the mean ± standard error (SEM) (n = 08). *: p < 0.001 vs Group I, and #: p < 0.05 vs Group II using non-parametric Kruskal-Wallis test.

Fig 5. Effect of Pistacia lentiscus essential oil on 7,12-dimethylbenz(a)anthracene, inducing antioxidant enzyme disorders in the mammary tissue and plasma of C57Bl/6 female mice.

All animals were pretreated with two doses of DMBA (20 and 50 mg/kg, b.w. dose) associated with progesterone and estrogens treatment until mammary tumor induction. Groupe I: had not received any treatment, while group II and III received PL EO (280 and 570 mg/kg, b.w. dose respectively). Results are represented by the mean ± standard error (SEM) (n = 08). *: p < 0.001 vs Group I, and #: p < 0.05 vs Group II using non-parametric Kruskal-Wallis test.