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Introduction: Colorectal cancer remains the third most common cancer diagnosis and fourth leading cause of cancer-related mortality worldwide. Purified cannabinoids have been reported to prevent proliferation, metastasis, and induce apoptosis in a variety of cancer cell types. However, the active compounds from Cannabis sativa flowers and their interactions remain elusive. Research Aim: This study was aimed to specify the cytotoxic effect of C. sativa-derived extracts on colon cancer cells and adenomatous polyps by identification of active compound(s) and characterization of their interaction. Materials and Methods: Ethanol extracts of C. sativa were analyzed by high-performance liquid chromatography and gas chromatograph/mass spectrometry and their cytotoxic activity was determined using alamarBlue-based assay (Resazurin) and tetrazolium dye-based assay (XTT) on cancer and normal colon cell lines and on dysplastic adenomatous polyp cells. Annexin V Assay and fluorescence-activated cell sorting (FACS) were used to determine apoptosis and cell cycle, and RNA sequencing was used to determine gene expression. Results: The unheated cannabis extracts (C2F), fraction 7 (F7), and fraction 3 (F3) had cytotoxic activity on colon cancer cells, but reduced activity on normal colon cell lines. Moreover, synergistic interaction was found between F7 and F3 and the latter contains mainly cannabigerolic acid. The F7 and F7+F3 cytotoxic activity involved cell apoptosis and cell cycle arrest in S or G0/G1 phases, respectively. RNA profiling identified 2283 differentially expressed genes in F7+F3 treatment, among them genes related to the Wnt signaling pathway and apoptosis-related genes. Moreover, F7, F3, and F7+F3 treatments induced cell death of polyp cells. Conclusions:C. sativa compounds interact synergistically for cytotoxic activity against colon cancer cells and induce cell cycle arrest, apoptotic cell death, and distinct gene expression. F3, F7, and F7+F3 are also active on adenomatous polyps, suggesting possible future therapeutic value.

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OBJECTIVES: The purpose of the present experimental research was to assess the efficacy of intraperitoneal administration of rosuvastatin in preventing the formation of postoperative peritoneal adhesions. MATERIAL AND METHOD: Peritoneal adhesions were induced in 120 male rats of the Wistar-Bratislava breed, divided into 4 groups (n=30), using a parietal and visceral abrasion model. Group I was designated as the control group; in the case of group II, a saline solution was administered intraperitoneally, while in the case of groups III and IV, rosuvastatin solution with a concentration of 10 mg/kg and 5 mg/kg, respectively, was administered in a single dose, during laparotomy. All rats were sacrificed on the 14th postoperative day and the peritoneal adhesions were assessed macroscopically and microscopically. RESULTS: The formation of peritoneal adhesions (assessed macroscopically by appreciating their number, the area covered, and the degree of severity) was statistically significantly more reduced in the subjects that received rosuvastatin (groups III and IV) as compared to the control group (p<0.05). The degree of fibrosis assessed microscopically was also statistically significantly reduced in groups III and IV as compared to the control group (p<0.05). CONCLUSIONS: Rosuvastatin administered intraperitoneally correlates with a reduction of peritoneal adhesions in rats. KEY WORDS: Rosuvastatin, Peritoneal adhesions, Prevention.

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OBJECTIVES: The purpose of this experimental study was to demonstrate the reduction of peritoneal adhesions formation in rats after intraperitoneal administration of rosuvastatin, due to its anti-inflammatory effect. METHOD: Peritoneal adhesions were induced in 120 Wistar-Bratislava rats divided into 4 groups (n=30), using a parietal and visceral (cecal) abrasion model. Group I was designated as control group; in group II, a saline solution was administered intraperitoneally; in groups III and IV, a single dose of rosuvastatin solution, 10 mg/kg and 5 mg/kg respectively, was injected intraperitoneally. The serum values of tumor necrosis factor (TNF-α) and interleukin-1 (IL-1α) were determined on day 1 and day 7 postoperatively (ELISA). Macroscopic assessment of the peritoneal adhesions was conducted on day 14. RESULTS: Rosuvastatin therapy induced a significant decrease of tumor necrosis factor serum levels in groups III and IV, on day 1 and day 7 (p<0.01). Intraperitoneal administration of rosuvastatin correlated with a decrease of mean interleukin-1α levels on postoperative day 1 in groups III (p=0.0013) and IV (p=0.00011), but not on day 7, where the differences were no longer statistically significant (p=0.8) The reduction of postoperative peritoneal adhesions in the experimental rat model is supported by the anti-inflammatory effect of rosuvastatin, mediated mainly by the tumor necrosis factor. CONCLUSIONS: Rosuvastatin prevents the formation of postoperative peritoneal adhesions in rats. This effect may be linked to the inhibition of proinflammatory cytokines release in the early stages of adhesions formation. The present study suggests that rosuvastatin may be an efficient pharmacological agent in the prevention of postoperative peritoneal adhesions development, and requires further studies as it has a promising application value.

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Introduction: Inflammatory bowel diseases (IBDs) include Crohn's disease, and ulcerative colitis. Cannabis sativa preparations have beneficial effects for IBD patients. However, C. sativa extracts contain hundreds of compounds. Although there is much knowledge of the activity of different cannabinoids and their receptor agonists or antagonists, the cytotoxic and anti-inflammatory activity of whole C. sativa extracts has never been characterized in detail with in vitro and ex vivo colon models. Material and Methods: The anti-inflammatory activity of C. sativa extracts was studied on three lines of epithelial cells and on colon tissue. C. sativa flowers were extracted with ethanol, enzyme-linked immunosorbent assay was used to determine the level of interleukin-8 in colon cells and tissue biopsies, chemical analysis was performed using high-performance liquid chromatography, mass spectrometry and nuclear magnetic resonance and gene expression was determined by quantitative real-time PCR. Results: The anti-inflammatory activity of Cannabis extracts derives from D9-tetrahydrocannabinolic acid (THCA) present in fraction 7 (F7) of the extract. However, all fractions of C. sativa at a certain combination of concentrations have a significant increased cytotoxic activity. GPR55 receptor antagonist significantly reduces the anti-inflammatory activity of F7, whereas cannabinoid type 2 receptor antagonist significantly increases HCT116 cell proliferation. Also, cannabidiol (CBD) shows dose dependent cytotoxic activity, whereas anti-inflammatory activity was found only for the low concentration of CBD, and in a bell-shaped rather than dose-dependent manner. Activity of the extract and active fraction was verified on colon tissues taken from IBD patients, and was shown to suppress cyclooxygenase-2 (COX2) and metalloproteinase-9 (MMP9) gene expression in both cell culture and colon tissue. Conclusions: It is suggested that the anti-inflammatory activity of Cannabis extracts on colon epithelial cells derives from a fraction of the extract that contains THCA, and is mediated, at least partially, via GPR55 receptor. The cytotoxic activity of the C. sativa extract was increased by combining all fractions at a certain combination of concentrations and was partially affected by CB2 receptor antagonist that increased cell proliferation. It is suggested that in a nonpsychoactive treatment for IBD, THCA should be used rather than CBD.

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MAIN CONCLUSION: MAX2/strigolactone signaling in the endodermis and/or quiescent center of the root is partially sufficient to exert changes in F-actin density and cellular trafficking in the root epidermis, and alter gene expression during plant response to low Pi conditions. Strigolactones (SLs) are a new group of plant hormones that regulate different developmental processes in the plant via MAX2, an F-box protein that interacts with their receptor. SLs and MAX2 are necessary for the marked increase in root-hair (RH) density in seedlings under conditions of phosphate (Pi) deprivation. This marked elevation was associated with an active reduction in actin-filament density and endosomal movement in root epidermal cells. Also, expression of MAX2 under the SCARECROW (SCR) promoter was sufficient to confer SL sensitivity in roots, suggesting that SL signaling pathways act through a root-specific, yet non-cell-autonomous regulatory mode of action. Here we show evidence for a non-cell autonomous signaling of SL/MAX2, originating from the root endodermis, and necessary for seedling response to conditions of Pi deprivation. SCR-derived expression of MAX2 in max2-1 mutant background promoted the root low Pi response, whereas supplementation of the synthetic SL GR24 to these SCR:MAX2 expressing lines further enhanced this response. Moreover, the SCR:MAX2 expression led to changes in actin density and endosome movement in epidermal cells and in TIR1 and PHO2 gene expression. These results demonstrate that MAX2 signaling in the endodermis and/or quiescent center is partially sufficient to exert changes in F-actin density and cellular trafficking in the epidermis, and alter gene expression under low Pi conditions.

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BACKGROUND AND AIMS: The present study conducted from March 2012 to July 2013 aimed to evaluate from echocardiographic point of view the effects of peripheral intravenous administration of mesenchymal stem cells (MSCs) in laboratory rabbits presenting 30 days old chronic myocardial infarction. MATERIAL AND METHODS: 30 days after the induction of an acute myocardial infarction in 40 laboratory rabbits by direct ligation of the left anterior descending coronary artery at about 10 mm from the apex, we injected 1×106 MSCs in the auricular vein in a group of 30 rabbits, and a group of 10 rabbits were used as controls. 30 days after the injection of stem cells the left ventricular (LV) fractional shortening (FS) was evaluated by echocardiography and compared with the control rabbits. RESULTS: In control rabbits, echocardiography revealed akinesis of apex, interventricular septum kinetics was also impaired, FS being approximately 6%. In 80% (24 rabbits) of the injected rabbits the FS of the LV was significantly greater than in the witness group (26+/-2%, p<0.0001). At 13.3% (4 rabbits) of the injected rabbits the FS of the LV showed no improvement in comparison with the control group (6.5+/-1%). CONCLUSION: An improvement of LV SF 30 days after MSCs were injected(p<0.0001) was noted. We have to further determine if this improvement of the LV function is correlated with any histopathological changes and if it is not lost in time. Also, further studies needs to evaluate if there is any significant change in the overall mortality.

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Bacterial soft rot caused by Pectobacterium carotovorum subsp. carotovorum (Pcc) is one of the most devastating diseases of Ornithogalum species. No effective control measures are currently available to use against this pathogen; thus, introduction of resistant genes via genetic transformation into this crop is a promising approach. Tachyplesin I, an antimicrobial peptide, has been shown to effectively control numerous pathogenic bacteria, including Pcc. In this study, liquid-grown cell clusters of Ornithogalum dubium and Ornithogalum thyrsoides were bombarded with a pCAMBIA2301 vector containing a celI leader sequence fused to a gene encoding tachyplesin I, a neomycin phosphotransferase (nptII) gene that served as a selectable marker and a ß-glucuronidase (GUS) gene that served as a reporter. Selection was carried out in the dark in liquid medium containing 80mg/L kanamycin. Regeneration was executed in the light after 6-14 months depending on the cultivar. Hundreds of transgenic plantlets were produced and their identity was confirmed through GUS activity assays. PCR and RT-PCR were used to confirm the presence of the target, reporter and selection genes in the divergent lines of plantlets. The resistance of the O. dubium plants to Pcc was evaluated in vitro, following infection with a highly virulent isolate from calla lily. Although control plantlets were completely macerated within a week, 87 putative transgenic subclones displayed varying levels of disease resistance. During three growing seasons in the greenhouse, the transgenic O. dubium lines grew poorly, whereas the transgenic O. thyrsoides plants grew similarly to non-transgenic plants.

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Dermatofibrosarcoma protuberans is a rare superficial tumor characterized by high rates of local recurrence and low risk of metastasis. Dermatofibrosarcoma protuberans occurs most commonly on the trunk and proximal extremities, it affects all races, and often develops between the second and the fifth decade of life. The tumor grows slowly, typically over years. We present a rare case of a young male patient, 21 years old, with an asymptomatic calf tumor which was suspected to be an angioma, but after the initial excision histology and imunohistochemistry proved to be a Dermatofibrosarcoma protuberans without safety limits. After 2 weeks, we excised the remaining scar with 4 cm tissue limit and the defect was covered using an adipofascial reversed sural flap from the posterior part of the left calf and after another 2 weeks we applied a skin graft from the thigh. The patient had a good evolution, with full recovery, without local recurrences or metastasis, and the histology was within good safety limits.

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BACKGROUND: Critical limb ischemia (CLI) is associated with an increased risk of limb amputation, low quality of life and cardiovascular death. The aim of this study is to identify the prognostic factors of mortality, revascularization failure and amputation failure, as part of risk factors for athero-sclerosis and comorbidities. PATIENTS AND METHODS: We examined 198 patients operated for CLI. Cox analysis was performed to discern the factors that were associated with failure of initial surgical therapy and death. RESULTS: For survival analysis, a significant model emerged with hypertension (p=0.00), cardiac comorbidities (p=0.00), renal comorbidities (p=0.04) and respiratory comorbidities (p=0.02) as significant predictors. Regarding the time to amputation failure, there was a significant model with insulin treated diabetes (p=0.00), coronary artery disease (p=0.02) and cerebrovascular disease (p=0.05) as significant predictors. CONCLUSIONS: Significant predictors for mortality in CLI patients are high risk hypertension, severe coronary artery disease, renal failure requiring dialysis and chronic obstructive pulmonary disease. The association of these prognostic factors results in a proportional decrease of survival. The predictors for amputation failure were, in addition to local factors, insulin treated diabetes, coronary artery disease and cerebrovascular disease. The revascularization for limb salvage depends on the correct indication and accurate surgical technique.

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INTRODUCTION: Rat mesenchymal stem cells (MSCs) represent a small portion of the cells in the stromal compartment of bone marrow and have the potential to differentiate into new blood vessel and other tissues. MSCs transplantation in tissues from critical limb ischemia model in rat may represent a therapeutic applications of vascular regeneration. AIM: The aim of this study was to isolate with a simple method the rat bone marrow stromal cells. Then the adherent cells were labeled with 5-bromo-2-deoxyuridine (BrdU, Sigma) and injected in the gastrocnemius and adductor muscle of ischemic hind limbs in order to demonstrate their presence in the critical limb ischemia model in rat. MATERIAL AND METHODS: MSCs were isolated from Wistar rats, 8 weeks of age. The MSCs were labeled in vitro for later identification by adding 10 µg/mL 5-bromo-2 -deoxyuridine (BrdU, Sigma). RESULTS: Small colonies of fibroblast-like cells were seen after several days of primary culture. These colonies increased in size and were subcultured after 15-18 days. CONCLUSION: The MSCs obtained in this study presented a stable undifferentiated phenotype under normal culture conditions. MSCs are easy to isolate, culture, and detect in in vivo culture. These cells are characterized by high plasticity and could have an important role in angiogenesis.

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INTRODUCTION: Chronic lower limb ischemia (CLLI) leads to endothelial cell dysfunctions and endothelial lesions. The use of substances that release nitric oxide and activate endothelial nitric oxide synthase has proved to be useful in increasing angiogenesis and arteriogenesis under critical ischemia conditions. OBJECTIVES: To investigate the therapeutic effect of Sildenafil and Donepezil with a vasodilating action in experimentally induced CLLI and on serum redox homeostasis. MATERIAL AND METHOD: The research was performed in 3 groups of rats (n=10 animals/group) with experimentally induced CLLI: group I - control group; group II - animals treated postoperatively with a therapeutic dose of sildenafil, and group III - animals treated postoperatively with a therapeutic dose of donepezil. Oxidative stress (OS) indicators (malondialdehyde - MDA, protein carbonyls - PC), antioxidant (AO) defense indicators (reduced glutathione - GSH and oxidized glutathione - GSSH), and ceruloplasmin (CP) were determined on days 7, 14, 21 and 30. Statistical processing was performed using the Excel application (Microsoft Office 2007), with the StatsDirect v.2.7.2 software. RESULTS: Changes in OS were evidenced in all groups on account of a decrease in MDA and PC. The greatest OS decrease in all groups was on day 30. AO defence changes were represented by decreased levels of GSH and GSSG in all groups, at the studied moments. Intracellular AO defense in the cytosol, nucleus and mitochondria was similar in all groups, (decreased GSH, GSSG and GSH/GSSG ratio). We found increased extracellular levels of GSH, GSSG, and CP and increased extracellular GSH/GSSG ratio at level compared to values on day 7. CONCLUSIONS: 1) The administration of sildenafil (group II) and donepezil (group III) has favorable effects on reducing OS in experimentally induced CLLI. 2) Sildenafil and Donepezil administration stimulates extracellular AO defense on account of CP. 3) Sildenafil and Donepezil administration influences intracellular redox homeostasis on account of the GSH/GSSG couple, the major redox buffer in the body.

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INTRODUCTION: Ischemic heart disease is a major public health problem in western countries. Appropriate animal experimental models of chronic myocardial infarction is an essential first step in order to investigate and develop new therapeutic interventions. AIM: The aim of this study was to find an optimal place for a coronary artery ligation to induce an optimal chronic myocardial infarction and also a new heart approach that will not require oro-tracheal intubation. MATERIAL AND METHODS: To achieve these goals we used a group of rabbits and after induction of anesthesia and cardiac exposure by rib osteotomy (rib III, IV and V) at the costo-sternal junction level on the right side we performed three different left anterior descending artery (LAD) ligation at different distances (5, 10 and 15 mm) in relation to the apex. Thirty days after the acute myocardial infarction, we correlated laboratory investigations (serology, ECG, cardiac ultrasound) with histopathological findings. RESULTS: Heart approach achieved by rib osteotomy (rib III, IV and V) at the costo-sternal junction level on the right side, maintains the integrity of the ribcage, allowing it to take part in respiratory movements and the animal model does not need oro-tracheal intubation. Ligation of LAD at 15 mm from the apex was incompatible with life; ligation of LAD at 5 mm from the apex does not achieved transmural myocardial infarction and ligation of LAD at 10 mm from the apex achieved a transmural myocardial infarction of the left ventricle which also involved the distal part of the interventricular septum. CONCLUSION: Ligation of LAD at 10 mm from the apex achieved a transmural myocardial infarction of the left ventricle, is in an easily accessible area from technical point of view, it is sufficiently expanded to induce hemodynamic effects that can be quantified with paraclinical examination and also it is compatible with the experimental animal life. If the heart is approached by rib III, IV and V osteotomy at the costo-sternal junction level on the right side combined with neuroleptic anaesthesia, the animal does not need assisted ventilation.