Impacts of some clinicopathodemography and colorectal tissues key cell cycle and mucin stabilizing molecules on the metastasis trend in colorectal cancer patients.

BACKGROUND: We aimed to evaluate the various clinicopathodemographical, epidemiological, and molecular contributors to cumulatively worldwide metastatic colorectal cancer (CRC) in CRC patients from a highly populated area in northeastern Iran to pinpoint metastasis risk. METHODS: A retrospective clinical material-based cohort including a total of 6260 registered CRC patients, of whom 3829 underwent surgery, from regional university hospitals, during 2006-2016, were analyzed for the clinicopathodemographical aspects of age, sex, stage of CRC, history of smoking, type 2 diabetes (T2D), hypertension, body mass index (BMI), familial/occupational status, post-surgery survival period and mRNA/protein expression of mucin stabilizer (B3GALNT2), mucin I (MUC1), key cell cycle molecules (i.e., P53 and Ki67), and MMR-related genes. Factors were set to estimate the risk of metastatic CRC and mortality. RESULTS: Predominant adenocarcinomatous CRCs were found in colon. Post-surgery survival period of metastatic CRC patients was remarkably longer in patients aged > 50 compared to those aged < 50 years, and worse in females than males. B3GALNT2high, MUChigh, P53low, and Ki67high mRNA/protein expression in the metastatic stage III CRC along with T2D and hypertension were associated with increased metastasis/mortality, with more worsening in males, older, BMI > 25, urban residing, and employed individuals, indicative of non-genetic attributable factors. CONCLUSION: B3GALNT2, MUC1, and "Ki67" can be used as promising biomarkers for prognosis and early diagnosis of increasingly/predominantly non-genetic/environmental originated metastatic CRCs.

The immunomodulatory effect of minocycline on gene expression of inflammation related cytokines in lipopolysaccharide-treated human peripheral blood mononuclear cells.

To evaluate the immunomodulatory effect of minocycline, the present study was carried out on the gene expression of toll-like receptor type-4 (TLR4) and some pro-inflammatory (IL-1ß, IL-6) and anti-inflammatory cytokines (IL-10) associated with lipopolysaccharide (LPS) -induced inflammation in human peripheral blood mononuclear cells (PBMCs). The PBMCs were collected and then 5.4 × 106 PBMCs/mL were used in eight groups as follows: control group (only media), LPS group (only LPS), methylprednisolone (Pred) group (LPS plus Pred), meloxicam (Melo) group (LPS plus Melo), three minocycline groups [M1, M5 and M25] (LPS plus 1, 5, and 25 µg/mL minocycline, respectively) and minocycline control (MC) group (5 µg/mL minocycline). After incubation for 24 h, the PBMCs were subjected to quantitative PCR assays. Gene expression levels of TLR4 were not changed in any groups. The IL-1ß levels were increased in the LPS group but the increases were much more intense in the other groups except Pred group. Compared with control group, IL-6 levels increased significantly in Melo, M1 and M25 groups. Significant increases of IL-10 levels were also observed in Melo, M25 and MC groups. It can be concluded that minocycline had dual pro- and anti-inflammatory activities with potential clinical immunomodulatory effects.

The anti-inflammatory effects of minocycline on lipopolysaccharide-induced paw oedema in rats: a histopathological and molecular study.

Minocycline is a semi-synthetic antimicrobial agent with claimed anti-inflammatory properties reported from different experimental models. This study was aimed to evaluate the anti-inflammatory effects of minocycline, compared to the actions of two common anti-inflammatory agents, on lipopolysaccharide (LPS)-induced paw oedema through some clinical, histopathological, haematological and molecular analyses. Forty-eight rats were divided into eight groups (n = 6). In control group (Ctrl), each animal was injected with normal saline into its sub-plantar region of hind paw. In groups 2-7, hind paw oedema was induced by injection of LPS. One hour before injections, groups 1 (Ctrl) and 2 (LPS) were treated orally with distilled water, 3 and 4 with methylprednisolone (Pred) and meloxicam (Melo) and 5-7 with minocycline in doses of 50, 150 and 450 mg/kg (M50, M150 and M450, respectively). The 8th group (MC) was given minocycline (150 mg/kg) orally and normal saline was injected into sub-plantar region. Paw swelling and body temperature were assessed at 0, 2, 4, 6 and 24 h post-injections. At 24 h, samples of blood and liver, kidney, spleen and hind paw tissues were taken for haematological and histopathological examinations. Some samples of the paw were also obtained for molecular analysis of some inflammatory-related cytokines at mRNA level. Paw swelling and body temperature increased in all LPS-injected groups 2 h post-injection. In LPS group, they remained significantly increased up to 24 h; however, these parameters decreased to normal in Pred, Melo and all minocycline groups. The histological findings showed mild-to-moderate signs of inflammation in tissue samples of groups 2-6, but not in group M450. Additionally, gene expression of pro-inflammatory cytokines (IL-1ß and IL-6) increased significantly in LPS group compared to other groups. In conclusion, this study supports the role of minocycline as an anti-inflammatory agent with effects comparable to those of meloxicam and methylprednisolone.

Naturally Occurring Level of Aflatoxin B1 Injures Human, Canine and Bovine Leukocytes Through ATP Depletion and Caspase Activation.

Aflatoxin (AF) B1 is a potent hepatotoxic, mutagenic, teratogenic mycotoxin and may cause immune suppression/dysregulation in humans and animals. Toxic effects of AFB1 on key mammalian immune cells (ie, leukocytes) needs to be mechanistically elucidated. In this study, along with the determination of AFB1's LC50 for certain leukocytes, we analyzed the effect of naturally occurring levels of AFB1 on apoptosis/necrosis of neutrophils, lymphocytes, and monocytes from healthy young humans (20- to 25-year-old male), dogs (1- to 2-year-old Persian/herd breed), and cattle (1- to 2-year-old cattle). Leukocytes were incubated for approximately 24 hours with naturally occurring levels of AFB1 (10 ng/mL). Intracellular adenosine triphosphate (ATP) depletion and caspase-3/7 activity were then determined by luciferase-dependent bioluminescence (BL). Furthermore, the necrotic leukocytes were measured using propidium iodide (PI)-related flow cytometry. A significant decrease (24%-45%, 33.2% ± 2.7%) in intracellular ATP content was observed in AFB1-treated neutrophils, lymphocytes, and monocytes in all studied mammals. Also, with such a low level (10 ng/mL) of AFB1, BL-based caspase-3/7 activity (BL intensity) in all 3 tested mammalian leukocyte lineages was noticeably increased (∼>2-fold). Flow cytometry-based PI staining (for viability assay) of the AFB1-treated leukocytes showed slightly/insignificantly more increase of necrotic (PI+) neutrophils, lymphocytes, and monocytes in human, dogs, and cattle. Even though in vitro LC50s for AFB1' (∼20,000-40,000 ng/mL) were approximately 2,000 to 4,000 times higher than background, these studies demonstrate leukocytes from human and farm/companion animals are sensitive to naturally occurring levels of AFB1. The observed in vitro ATP depletion and caspase activation in AFB1-exposed leukocytes can partially explain the underlying mechanisms of AFB1-induced immune disorders in mammals.

Human Microglial Cells Undergo Proapoptotic Induction and Inflammatory Activation upon in vitro Exposure to a Naturally Occurring Level of Aflatoxin B1.

OBJECTIVE: Knowledge regarding interactions of AFB1 with the human nervous system and how a naturally occurring level of AFB1 could potentially induce neuroimmune dysregulation is very limited. To assess the cellular effects of AFB1 on the human brain, we used the human microglia cell line CHME5 as a model to pinpoint its potential in vivo translation. METHODS: We used the CHME5 cell line culture system, multiplex qPCR, (chemi)bioluminescence, Luminex ELISA, and flow cytometry assays to evaluate the toxic effects of a naturally occurring level of AFB1 on human microglia. RESULTS: A low concentration of AFB1 upregulates the mRNA expression of many proinflammatory molecules, such as TLRs, MyD88, NFκB, and CxCr4, induces intracellular ATP depletion, and increases caspase-3/7 activity at different time points following exposure to the toxin. Furthermore, AFB1-exposed microglia secreted significantly higher levels of IFN-γ and GM-CSF after treatment. We also observed a slight increase in the percentage of apoptotic microglia (annexin V+/PI-) at 48 h posttreatment. CONCLUSION: Our work confirmed that the environmentally relevant level of AFB1 could cause an inflammatory reaction in human microglial cells that is potentially harmful or toxic to the homeostasis of the human central nervous system and might increase susceptibility to neurodegenerative diseases.

High soil and groundwater arsenic levels induce high body arsenic loads, health risk and potential anemia for inhabitants of northeastern Iran.

Arsenic bioavailability in rock, soil and water resources is notoriously hazardous. Geogenic arsenic enters the body and adversely affects many biochemical processes in animals and humans, posing risk to public health. Chelpu is located in NE Iran, where realgar, orpiment and pyrite mineralization is the source of arsenic in the macroenvironment. Using cluster random sampling strategy eight rocks, 23 soils, 12 drinking water resources, 36 human urine and hair samples and 15 adult sheep urine and wool samples in several large-scale herds in the area were randomly taken for quantification of arsenic in rock/soil/water, wool/hair/urine. Arsenic levels in rock/soil/water and wool/hair/urine were measured using inductively coupled plasma spectroscopy and atomic absorption spectrophotometry, respectively. While arsenic levels in rocks, soils and water resources hazardously ranged 9.40-25,873.3 mg kg(-1), 7.10-1448.80 mg kg(-1) and 12-606 µg L(-1), respectively, arsenic concentrations in humans' hair and urine and sheep's wool and urine varied from 0.37-1.37 µg g(-1) and 9-271.4 µg L(-1) and 0.3-3.11 µg g(-1) and 29.1-1015 µg L(-1), respectively. Local sheep and human were widely sick and slightly anemic. Hematological examination of the inhabitants revealed that geogenic arsenic could harm blood cells, potentially resulting in many other hematoimmunological disorders including cancer. The findings warn widespread exposure of animals and human in this agroecologically and geopolitically important region (i.e., its proximity with Afghanistan, Pakistan and Turkmenistan) and give a clue on how arsenic could induce infectious and non-infectious diseases in highly exposed human/animals.

Seroepidemiology of Q fever in one-humped camel population in northeast Iran.

Coxiella burnetii, an obligate intracellular bacterium, is the causative agent of important zoonotic Q fever. It is the etiological agent of coxiellosis or Q fever in animals and human. This seroepidemiological survey was conducted to determine the seroprevalence of coxiellosis in increasingly camel raised population in vast area of Khorasan (North, South, and Razavi) provinces, northeast Iran. Using cluster random sampling strategy, 167 camels in 11 counties were selected as serum samples. Sera were assayed for antibody against C. burnetii using a Q fever ELISA kit. Logistic regression model was used to insight the contributing risk factor(s) of Q fever in the study area. C. burnetii was widely distributed throughout the study area. Seroprevalence of C. burnetii at animal level was 28.7 % [(95 % confidence interval (CI): 21.83, 35.56)] for camel populations. The proportion of seropositivity for camels in the studied counties ranged from 0 to 63.6 %. Logistic regression model showed that age correlated with seroprevalence of coxiellosis at the individual level in camels (P < 0.05). This study showed that a relatively high proportion of camels are seropositive to C. burnetii. Considering the economic, zoonotic, and public health importance of Q fever, percussion measures are to be implemented to prevent spreading of C. burnetii and zeroing the risk of Q fever in farm animals and human in this agro-ecologically and geopolitically important region.

Cytochrome P450 isoforms are differently up-regulated in aflatoxin B1-exposed human lymphocytes and monocytes.

CONTEXT: Aflatoxins (AFs) are highly hazardous mycotoxins with potent carcinogenic, mutagenic and immune disregulatory properties. Cytochrome P450 (CYP) isoforms are central for enhanced AFB1 toxicity in situ. It remains to be seen whether and how these AFB1 activators work in human leukocytes. OBJECTIVE: To investigate the involvement of CYP isoforms in AFB1 toxicity of circulating mononuclear cells, we examined the impact of environmentally relevant levels of AFB1 on lymphocytes and monocytes. MATERIALS AND METHODS: Very low and moderate doses of AFB1 with/without CYP inducers on transcription of key CYP isoforms and toll-like receptor 4 (TLR4) were examined in human lymphocytes, monocytes and HepG2 cells; cell cycle distribution and viability were also analyzed in AFB1-exposed lymphocytes and monocytes. RESULTS: Only CYP1A1, CYP1B1, CYP3A4, CYP3A5 and CYP3A7 expressed in lymphocytes and monocytes. TLR4 much more expressed in monocytes than in lymphocytes, but HepG2 showed little TLR4 transcription. While CYP1A1, CYP1B1 and CYP3A4 were highly induced by AFB1 in monocytes, in lymphocytes only CYP1A1 was induced. Among CYP1A1, CYP1B1 and CYP3A4 only CYP1A1 responded to low and moderate levels of AFB1. Enhanced transcripts of CYPs by AFB1 yielded little synergies on TLR4 transcription in lymphocytes and monocytes. Cell cycle arrest and necrosis were also detected in AFB1-exposed lymphocytes and monocytes. CONCLUSIONS: Our novel findings indicate that AFB1 more intensively stimulates CYP genes expression in monocytes than in lymphocytes. Mechanistically, this could explain a more pronounced immunotoxicity of AFB1 in myeloid than in lymphoid lineage cells in vitro/situ/vivo.

Environmentally Relevant Level of Aflatoxin B1 Dysregulates Human Dendritic Cells Through Signaling on Key Toll-Like Receptors.

Aflatoxins (AFs) are highly hazardous fungal biometabolites usually present in feeds and foods. Aflatoxin B1 (AFB1) is the most toxic and a known carcinogen. Toll-like receptors (TLRs), highly expressed by myeloid dendritic cells (DC), are key innate immune-surveillance molecules. Toll-like receptors not only sense pathogen-associated molecular patterns but also contribute to infections and cancer. To assess AFB1-TLR interactions on human myeloid DC, pure CD11c+ DC were generated from monocytes isolated from healthy individuals and then exposed to relevant level of AFB1 for 2 hours. Both quantitative polymerase chain reaction and flow cytometric assays were used to quantify, respectively, expression of TLR2 and TLR4 at the messenger RNA (mRNA) and protein levels in these DC. Levels of interleukin (IL) 1ß, IL-6, and IL-10 were also analyzed in AFB1- and mock-treated DC. Compared to nontreated CD11c+ DC, expression levels of both TLR2 and TLR4 mRNA and proteins were significantly upregulated in AFB1-treated cells. Further, although IL-10 levels in AFB1-treated DC were similar to those in the mock-treated DC, the AFB1-exposed DC secreted higher amounts of IL-1ß and IL-6. Dendritic cells are sensitive to environmentally relevant level of AFB1, and TLR2 and TLR4 are involved in sensing AFB1 Considering the broad roles of TLR2, TLR4, and DC in immunity and infections, our novel findings open a new door to understanding the molecular mechanisms and functional consequences of AFB1 in inducing immunodysregulation, immunotoxicity, and thus (non)infectious diseases in humans.

Biologically relevant doses of mixed aflatoxins B and G up-regulate MyD88, TLR2, TLR4 and CD14 transcripts in human PBMCs.

CONTEXT: Aflatoxins (AFs) are highly hazardous carcinogenic mycotoxins originated from very common fungi present in the environment. Their effect on key immune-surveillance molecules is unclear. OBJECTIVE: We aimed to examine the effect of mixed AFs on immunologically relevant molecules and on viability in human peripheral blood mononuclear cells (PBMCs), in conditions similar to those occurring naturally, i.e. using a mixture of environmentally relevant levels of AFB1, AFB2, AFG1 and AFG2. MATERIALS AND METHODS: We evaluated the mRNA expression of MyD88, toll-like receptor (TLR)-2, TLR4 and CD14, in human PBMCs treated with a mixture of AFB1, AFB2, AFG1 and AFG2 at different doses for 2, 12 and 24 h. We used qRT-PCR to assess changes in transcripts of MyD88, TLR2, TLR4 and CD14 in PBMCs. We also evaluated the viability of PBMCs exposed to AFs. RESULTS: Biologically relevant levels of mixed AFs elicited early immune modulation in human PBMCs. qRT-PCR results showed several folds increase of MyD88, TLR2, TLR4 and CD14 transcripts in PBMCs as early as 2 h post-exposure to mixed AFs. Kinetics and dose-response of the up-regulation differed for mentioned gene transcripts. Further, prolonged exposure to mixed AFs decreased PBMCs viability. CONCLUSION: Immunotoxicity of AFs on PBMCs may be mediated by up-regulation of key immune-surveillance molecule transcripts. The description of these effects induced by AFs on PBMCs are novel and should be taken into account when considering AF-related infectious and noninfectious diseases in areas highly exposed to AFs.

Anti-inflammation-based treatment of atherosclerosis using Gliclazide-loaded biomimetic nanoghosts.

In the study, a biomimetic platform for anti-inflammatory-based treatment of atherosclerotic plaque was developed. Gliclazide (GL) as an anti-inflammasome agent was encapsulated in PLGA nanoparticles (NP), which were coated by monocyte membrane using an extrusion procedure. The size and zeta potential of the nanoghost (NG) changed to 292 and - 10 nm from 189.5 to -34.1 in the core NP. In addition, the actual size of 62.5 nm with a coating layer of 5 nm was measured using TEM. The NG was also showed a sustained release profile with the drug loading content of about 4.7%. Beside to attenuated TNFα, decrease in gene expression levels of NLRP3, MyD88, NOS, IL-1ß, IL-18 and caspases 1/3/8/9 in LPS-primed monocytes exposed to NG strongly indicated remarkable inflammation control. After systemic toxicity evaluation and pharmacokinetic analysis of NP and NG, intravenous NG treatment of rabbits with experimentally induced atherosclerosis revealed remarkably less plaque lesions, foam cells, lipid-laden macrophages, and pathological issues in tunica media of aorta sections. Higher expression of CD163 than CD68 in aorta of NG-treated rabbits strongly reveals higher M2/M1 macrophage polarization. The bio/hemocompatible, biomimetic and anti-inflammatory NG can be considered as a potential platform for immunotherapy of particularly atherosclerosis in the field of personalized medicine.

Noticeable immune dysregulation-and-suppression in parvovirus affected dogs.

Canine parvovirus type 2 (CPV-2) is one of the most common causes of infectious diarrhea in small animals, with high mortality and morbidity. Information on the specific treatment option(s) for CPV diseases (CPVD) is unachievably little. So, the treatment is mainly supportive one. Disruption of dog's innate immune system in viral diseases simply occurs; presumably, the CPV-2 may change the level of some TLRs, interleukins, CD4 and CD8 in the leukocytes of CPVD dogs, and disruptive activities of these immune molecules might be attributable to severe CPVD in dogs. Study on the role of the key immune molecules in CPVD is rare. Herein, by conducting and relating the clinical, para-clinical, immunological and molecular diagnostic tests, we tried to establish how some key immune molecules behave in blood of parvovirus affected dogs. As such, in the 1st study, the mRNA levels of TLR2, TLR4, TLR9, IL-1ß, IL-6, CD4 and CD8 genes in the leukocytes of CPVD were assessed with quantitative (q)RT-PCR along with CPV-2 detection by rapid immunochromatography and PCR tests. In a 2nd study, the same measurements as in the 1st study were evaluated in two groups of mild versus severe clinical signs of CPVD. Both in the 1st and the 2nd studies leukopenia, much more pronounced in the severe CPVD, and immune dysregulation were observed. In the 1st study, a noticeable increase in the mRNA levels of TLR2 and TLR4 was detected with a slight decrease in TLR9 and a significant decrease in the expression of IL-1ß, IL-6, CD4 and CD8 in leukocytes of CPV-infected dogs. Compared to the mild CPVD, the intense of downregulating effects on those immune molecules in the 2nd study was remarkably much more pronounced in the severe CPVD. Overall, it proves strong immune dysregulation and suppression/incompetence and potential T-cells exhaustion in severely CPV-2-affected dogs. Technically and clinically, this would be substantially applicable in canine medicine. By targeting those key immune molecules and their signaling pathways, new clinicodiagnostic approaches for CPVD can be evolved, and biotechnicoclinically this would be substantially applicable in all physiopathological conditions of dogs.

Superparamagnetic Iron Oxide Nanoparticles Induce Apoptosis in HT-29 Cells by Stimulating Oxidative Stress and Damaging DNA.

Nanoparticles have garnered considerable scientific attention in recent years due to their diagnostic and therapeutic applications in cancer. The purpose of this study was to determine the effect of superparamagnetic iron oxide nanoparticles (Fe3O4 MNPs) on the induction of apoptosis in human colorectal adenocarcinoma cell line (HT-29) cells. The purpose of this study was to elucidate the mechanisms of apoptosis induced by Fe3O4 MNPs following MTT assay and to determine the optimal dose of 2.5 g/mL for inducing apoptosis in HT-29 cells. In HT-29 cells, Fe3O4 MNPs increased reactive oxygen species (ROS), calcium ion (Ca2+), and DNA damage. Additionally, the Fe3O4 MNPs significantly increased caspase 3 and 9 expression and decreased Bcl-2 expression at the protein and mRNA levels when compared to the control group (P = 0.0001). Fe3O4 MNPs also induced apoptosis in cancer cells by increasing the level of (ROS) and intracellular Ca2+, followed by an increase in caspase 3 and 9 expression and a decrease in Bcl-2 expression and direct DNA damage. Fe3O4 MNPs are an appropriate choice for colon cancer treatment based on their cell toxicity and induction of apoptosis in HT29 cells.

Study of the regulatory elements of the Ovalbumin gene promoter using CRISPR technology in chicken cells.

BACKGROUND: Hormone-dependent promoters are very efficient in transgene expression. Plasmid-based reporter assays have identified regulatory sequences of the Ovalbumin promoter that are involved in response to estrogen and have shown that the deletion of the steroid-dependent regulatory element (SDRE) and negative regulatory element (NRE) leads to a steroid-independent expression of a reporter. However, the functional roles of these regulatory elements within the native genomic context of the Ovalbumin promoter have not been evaluated. RESULTS: In this study, we show that the negative effects of the NRE element on the Ovalbumin gene can be counteracted by CRISPR interference. We also show that the CRISPR-mediated deletion of SDRE and NRE promoter elements in a non-oviduct cell can lead to the significant expression of the Ovalbumin gene. In addition, the targeted knock-in of a transgene reporter in the Ovalbumin coding region and its expression confirms that the truncated promoter of the Ovalbumin gene can be efficiently used for an estrogen-independent expression of a foreign gene. CONCLUSIONS: The methodology applied in this paper allowed the study of promoter regulatory sequences in their native nuclear organization.

Metabolic Disruption by Naturally Occurring Mycotoxins in Circulation: A Focus on Vascular and Bone Homeostasis Dysfunction.

Naturally occurring food/feed contaminants have become a significant global issue due to animal and human health implications. Despite risk assessments and legislation setpoints on the mycotoxins' levels, exposure to lower amounts occurs, and it might affect cell homeostasis. However, the inflammatory consequences of this possible everyday exposure to toxins on the vascular microenvironment and arterial dysfunction are unexplored in detail. Circulation is the most accessible path for food-borne toxins, and the consequent metabolic and immune shifts affect systemic health, both on vascular apparatus and bone homeostasis. Their oxidative nature makes mycotoxins a plausible underlying source of low-level toxicity in the bone marrow microenvironment and arterial dysfunction. Mycotoxins could also influence the function of cardiomyocytes with possible injury to the heart. Co-occurrence of mycotoxins can modulate the metabolic pathways favoring osteoblast dysfunction and bone health losses. This review provides a novel insight into understanding the complex events of coexposure to mixed (low levels) mycotoxicosis and subsequent metabolic/immune disruptions contributing to chronic alterations in circulation.

Long Term Oral Administration of Oregano Essence Effectively Relieves Polycystic Ovarian Rats through Endocrine and Inflammatory Balance.

Polycystic ovarian syndrome (PCOS) is alarmingly rising and sustainable therapy/prevention is needed. Here, we evaluated the therapeutic effects of oregano or Origanum vulgare (O. vulgare) essence (OE) on the PCOS rat model system. Vaginal smears monitored the estrous cycle of 40 virgin adult rats, and they received 2 mg estradiol valerate (EV)/0.2 ml corn oil intramuscularly to induce PCOS. At 60 days post-EV injection, all rats were evaluated for follicular development/cysts. The EV-induced PCOS rats were orally administered 250 and 500 mg/kgBW/day of OE for 30 days. OE was also further assessed for its predominant components along with hormonal, histological, and inflammatory-related gene expressions in the ovaries. The main components of the OE were predominantly pulegone (36.3), L-menthone (31.3%), far less piperitone (7.8%), isopiperitone (6.4%), isomenthol (3.6%), humulene epoxide II (2.2%), α-pinene (1.7%), and thymol (1.5%). Hormonal, histological, and inflammatory-related gene expression results showed >4-fold and 1.5-fold increase in FSH and progesterone; ∼50%, 85%, 45%, 55%, and 30% decreased in LH, estradiol, estrogen, testosterone, and AMH; and dose-dependently decreased in mRNA expression of IL-6, IL-1α, NF-kB, TNF-α, and IL-1ß by 25-65%, 55-75%, 15-40%, 30-55%, and 35-55%, respectively, and thus decreased the severity of PCOS, boosted endocrine balance, restored functional follicles and corpus luteum, and thus ovulation in PCOS rats. Overall, in the disrupted PCOS rats, OE oral treatment effectively relieved estradiol-induced PCOS rats via: (1) its endocrine balancing on GnRH, FSH, and LH and (2) its anti-inflammatory and antioxidant properties on ovary caused by OE's useful compounds like pulegone, thymol, and L-menthone. Though many aspects of the effects remain to be tested, such an underlying mechanistic reproductive regulatory effect observed in OE-administered rats further proves its sensible pharmaceutical applications in reproductive medicine and more specifically, PCOS.

The Effect of Thymus vulgaris on Hepatic Enzymes Activity and Apoptosis-Related Gene Expression in Streptozotocin-Induced Diabetic Rats.

Many diseases, including diabetes, are involved in the development of liver disorders through changes in the expression of genes such as apoptosis-related genes. In the present study, the effect of Thymus vulgaris (T. vulgaris) on hepatic enzyme activity and apoptosis-related gene expression in streptozotocin (STZ)-induced diabetic rats was examined. In this study, 50 adult male Wistar rats weighing approximately 200-220 g were divided into five groups. Diabetes was induced by an intraperitoneal injection of STZ (60 mg/kg). Following 18 days, all the animals in different groups were weighed and blood samples were taken from their cardiac veins. Gas chromatography-mass spectrometry (GC-MS) analysis revealed 45 different compounds in the T. vulgaris, including thymol (39.1%), p-cymene (20.63%), and γ-terpinene (14.85%). The results showed a significant increase in liver enzymes (aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP)) in diabetic or STZ mice compared to the control group (healthy mice) (P < 0.0001). The levels of AST, ALT, and ALP in rats treated with 200 mg/kg and 400 mg/kg of T. vulgaris extract showed a significant decrease in these enzymes in comparison with diabetic rats (P < 0.0001). The expression of caspase 3 and 9 genes in the groups treated with thyme significantly decreased compared to diabetic mice (P < 0.0001), and the expression of B-cell lymphoma-2 (Bcl-2) in the group receiving 400 mg/kg of thyme significantly increased compared to diabetic mice (P < 0.0001). Due to its antioxidant compounds, thyme improves the liver tissue cells in STZ-induced diabetic mice by reducing caspases 3 and 9 as well as increasing Bcl-2.

Neuroimmune disruptions from naturally occurring levels of mycotoxins.

Substantial pieces of evidence support the potential of exogenous toxins in disrupting neuroimmune homeostasis. It appears that mycotoxins are one of the noticeable sources of naturally occurring substances dysregulating the immune system, which involves the physiology of many organs, such as the central nervous system (CNS). The induction of inflammatory responses in microglial cells and astrocytes, the CNS resident cells with immunological characteristics, could interrupt the hemostasis upon even with low-level exposure to mycotoxins. The inevitable widespread occurrence of a low level of mycotoxins in foods and feed is likely increasing worldwide, predisposing individuals to potential neuroimmunological dysregulations. This paper reviews the current understanding of mycotoxins' neuro-immunotoxic features under low-dose exposure and the possible ways for detoxification and clearance as a perspective.

The effect of equine bone marrow-derived mesenchymal stem cells on the expression of apoptotic genes in neutrophils.

BACKGROUND: Bone marrow mesenchymal stem cells (BM-MSCs), as multipotent cells with self-renewal and plastic-adherent properties, have immunomodulatory effects on immune cells, including neutrophils. These cells are in close proximity in bone marrow (BM) sinusoids with non-multiplicative immature neutrophils. BM-MSCs exert their immunomodulatory effects on adjacent cells both directly (cell-to-cell contact) and indirectly (secretion of soluble factors). OBJECTIVES: The aim of this study was to evaluate the effect of equine bone marrow mesenchymal stem cells (BM-MSCs) on the expression of some pro- and anti-apoptotic genes (p53, survivin and Bcl2 ) in neutrophils co-cultured with BM-MSCs. METHODS: For this purpose, peripheral blood neutrophils were isolated and separately co-cultured for 12 hr with both BM-MSCs and the BM-MSCs΄ supernatant. Four groups were included: neutrophils with only culture media (as control), neutrophils co-cultured with BM-MScs, neutrophils cultured with BM-MSCs' supernatant and neutrophils cultured with lipopolysaccharide (LPS, as positive control). Then, the expression of mentioned genes (p53, survivin and Bcl2 ) was evaluated by quantitative polymerase chain reaction (qPCR). RESULTS: Compared with control neutrophils, in neutrophils co-cultured with both BM-MSCs and BM-MSCs' supernatant, the mRNA expression levels of p53, as pro-apoptotic gene, and survivin and Bcl2 , as anti-apoptotic genes, were remarkably increased and decreased (p < .05), respectively. CONCLUSIONS: These data revealed the notion that the direct contact of BM-MSCs is not obligatory for their effects on the apoptotic status of neutrophils and they affect neutrophils via soluble secreted factors, which is promising for clinical implications in equine medicine.

Apoptosis Induced by Ziziphora tenuior Essential Oil in Human Colorectal Cancer Cells.

Ziziphora (Cacotti in Persian) belongs to the Lamiaceae family (mint group) and is vastly found in Iran and Asia. This traditional medicinal plant is normally used as analgesic and for treatment of particular gastrointestinal diseases. Since colorectal cancer is one of the most common causes of death in the world and the second leading cause of cancer death among adults, there is a pressing need to inhibit this malignancy by using methods with minimal side effects. One of these methods is the use of natural resources such as medical plants. This study is aimed at investigating the expression of apoptosis-related genes in the adjacent culture of colorectal cancer epithelial cells (HT-29) with Ziziphora essential oil (ZEO). The essential oil was extracted from Ziziphora leaves, and its compounds were determined and then added to the HT-29 culture medium at different concentrations. After 24 hours, the HT-29 cells were harvested from the medium and cytotoxicity was analyzed by MTT assay. After MTT assay and determination of the percentage of apoptosis by flow cytometry, RNA extraction was performed and the expression levels of Bax, Bcl-2, caspase 3 (C3), and caspase 9 (C9) were analyzed using newly designed primers by reverse transcription (RT) qPCR method and GeniX6 software. Also, specific antibodies were used for western blot analyses of those molecules. GC analysis revealed 42 different compounds in the ZEO, including pulegone (26.65%), menthone (5.74%), thymol (5.51%), and menthol (1.02%). MTT assay showed that the concentration of 200 µg/ml of ZEO had the highest HT-29 cell death during 24 hours. After incubation with the concentration of 50 µg/ml of ZEO for 24 and 48 hours, caspase 3 and 9 gene expressions in the treated group increased compared to those in the control group (P < 0.001), while the Bcl-2 expression decreased. The results showed that having anticancer compounds, ZEO can increase C3 and C9 and decrease Bcl-2 expressions, causing apoptosis in HT-29 cells in vitro. This can lead to the use of ZEO as a factor for colorectal cancer treatment.