Isolation of alkaloids and anti-tumor activity of the crude methanolic extract of algerian Cytisus purgans

In this study, two known quinolizidine alkaloids which are sparteine and lupanine were isolated from the methanolic extract of the plant Cytisus purgans of Algerian flora by open column chromatography. These two compounds were identified on the basis of their spectral data (GC/MS, IR, MS, 1H and 13C). The anti-tumor activity of the crude methanolic extract of the aerial parts of the plant was also evaluated invitro against human breast cancer (MDA-MB-231) and human lung cancer (A549) cell lines using MTT assay

The effects of estradiol and human chorionic gonadotropin on acth cells in peripubertal female rats: A histological and stereological study

The effects of estradiol (EDP) and human chorionic gonadotropin (HCG) on histological and stereological features of adrenocorticotropic (ACTH) cells in peripubertal female rats were examined. The first group of females received five injections of EDP (0.25 mg/kg b.w.), every second day from the 4th to 14th day after birth, and was killed at the peripubertal stage. The second group of females was given two injections of pregnyl-gonadotrophinum chorionicum (HCG; 50 IU/kg body weight) on the 36th and 37th days after birth, and it was killed 24 h after the last treatment. The controls were injected with an equivalent volume of the corresponding vehicle. ACTH cells were immunohistochemically labeled and stereologically evaluated. Stereological analysis showed that the volume of ACTH cells and their volume density in peripubertal females treated with EDP, were decreased by 15.6% and 53.8% (p<0.05), respectively, compared to the controls. In HCG-treated animals, the observed parameters were increased by 39.2% and 15.4% (p<0.05), respectively, in comparison with the control females. These findings suggest that the application of EDP or HCG exerted opposite effects on the stereological features of pituitary ACTH cells

Carbon ions of different linear energy transfer (LET) values induce apoptosis & G2 cell cycle arrest in radio-resistant melanoma cells

© 2016, Indian Council of Medical Research. All rights reserved. Background & objectives: The main goal when treating malignancies with radiation is to deprive tumour cells of their reproductive potential. One approach is to induce tumour cell apoptosis. This study was conducted to evaluate the ability of carbon ions (12C) to induce apoptosis and cell cycle arrest in human HTB140 melanoma cells. Methods: In this in vitro study, human melanoma HTB140 cells were irradiated with the 62 MeV/n carbon (12C) ion beam, having two different linear energy transfer (LET) values: 197 and 382 keV/μm. The dose range was 2 to 16 Gy. Cell viability was estimated by the sulforhodamine B assay seven days after irradiation. The cell cycle and apoptosis were evaluated 48 h after irradiation using flow cytometry. At the same time point, protein and gene expression of apoptotic regulators were estimated using the Western blot and q-PCR methods, respectively. Results: Cell viability experiments indicated strong anti-tumour effects of12C ions. The analysis of cell cycle showed that12C ions blocked HTB140 cells in G2 phase and induced the dose dependent increase of apoptosis. The maximum value of 21.8 per cent was attained after irradiation with LET of 197 keV/μm at the dose level of 16 Gy. Pro-apoptotic effects of12C ions were confirmed by changes of key apoptotic molecules: the p53, Bax, Bcl-2, poly ADP ribose polymerase (PARP) as well as nuclear factor kappa B (NFκB). At the level of protein expression, the results indicated significant increases of p53, NFκB and Bax/Bcl-2 ratio and PARP cleavage. The Bax/Bcl-2 mRNA ratio was also increased, while no change was detected in the level of NFκB mRNA. Interpretation & conclusions: The present results indicated that anti-tumour effects of12C ions in human melanoma HTB140 cells were accomplished through induction of the mitochondrial apoptotic pathway as well as G2 arrest

Early effects of gamma rays and protons on human melanoma cell viability and morphology

The effects of irradiation with gamma rays and protons on HTB140 human melanoma cell morphology and viability were analyzed. Exponentially growing cells were irradiated close to the Bragg peak maximum of the 62-MeV proton beam, as well as with (60)Co gamma rays, with doses ranging from 8 to 24 Gy. The overall cell morphology was unchanged 6 and 48 h after gamma irradiation, also showing a relatively weak cell-inactivation level. After exposure to proton beam, considerable changes in cell morphology followed by stronger cell inactivation were achieved. Proliferation capacity of irradiated cells significantly decreased in both experimental set-ups. Higher ionization level of protons with respect to gamma rays, representing the main physical difference between these two types of radiation, was also revealed on the cell membrane level through larger pro-apoptotic capacity of protons

Formalin Fixation of Human Healthy Autopsied Tissues: The Influence of Type of Tissue, Temperature and Incubation Time on the Quality of Isolated DNA

Formalin fixation is a widely used method in histopathology that has certain limits. Formalin often leads to the degradation of DNA molecules in cancer tissues, which makes tissues unusable for molecular analysis. The other factors may also affect the quality of DNA isolated from fixed tissues. The aim of this study is to determine the impact of the incubation time and temperature on the quality of DNA molecules isolated from various healthy human tissues. The brain, lung and kidney tissues, excluded during the forensic autopsies of people who died of violent death, were fixed in phosphate-buffered formalin from 24h to two months. After the completion of the incubation period, the DNA was isolated using phenol-chloroform-isoamyl alcohol extraction method and the concentration and purity of the samples were determined spectrophotometrically. The degree of degradation of DNA was assessed by PCR reaction, by amplification of gene fragments which lengths were 150bp (GPD1) and 262bp (β-actin). The highest concentration, purity and preserved integrity of DNA were obtained from the brain samples. With prolonged tissue incubation times in formalin, the concentration and integrity of DNA decreased in all tissue samples, especially in the brain tissue, while the purity of DNA remained unchanged. Also, tissue fixation at +4°C contributed to a better quality of isolated DNA compared to DNA isolated from tissue fixed at room temperature. We can conclude that the type of human healthy tissue, temperature and the incubation time of formalin fixation have important influence on the concentration, purity and integrity of DNA during fixation of tissues excluded in the course of forensic autopsy

Thrombotic thrombocytopenic purpura: Etiopathogenesis, diagnostics and basic principles of treatment

© 2017, University of Kragujevac, Faculty of Science. All rights reserved. Thrombotic thrombocytopenic purpura (TTP) is a clinical syndrome that manifests with thrombocytopenia, microangiopathic haemolytic anaemia and symptoms and signs of kidney and brain damage, but it rarely involves other organs. The main pathophysiological cause of TTP is diminished metalloproteinase ADAMTS13 activity; the main function of ADAMTS13 is to degrade large multimers of the von Willebrand factor. Diminished activity of ADAMTS13 is caused either by a genetic mutation in the gene that codes ADAMTS13 (congenital TTP) or by antibodies that block ADAMTS13 enzyme activity or accelerate the degradation of ADAMTS13 (acquired TTP). Clinically, TTP presents most frequently with signs and symptoms of brain and kidney damage with concomitant haemorrhagic syndrome. TTP is suspected when a patient presents with a low platelet count, microangiopathic haemolytic anaemia (negative Coombs tests, low haptoglobine concentration, increased serum concentration of indirect bilirubin and lactate dehydrogenase, increased number of schysocytes in peripheral blood) and the typical clinical presentation. A definitive diagnose can be made only by measuring the ADAMTS13 activity. The differential diagnosis in such cases includes both typical and atypical haemolytic uremic syndrome, disseminated intravascular coagulation, HELLP syndrome in pregnant women and other thrombotic microangiopathies. The first line therapy for TTP is plasma exchange. In patients with acquired TTP, in addition to plasma exchange, immunosuppressive medications are used (corticosteroids and rituximab). In patients with hereditary TTP, the administration of fresh frozen plasma is sometimes required

Forensic genetics and genotyping

© 2019, University of Kragujevac, Faculty of Science. All rights reserved. Forensic genetics represents a combination of molecular and population genetics. Personal identification and kinship analysis (e.g. paternity testing) are the two main subjects of forensic DNA analysis. Biological specimens from which DNA is isolated are blood, semen, saliva, tissues, bones, teeth, hairs. Genotyping has become a basis in the characterization of forensic biological evidence. It is performed using a variety of genetic markers, which are divided into two large groups: bi-allelic (single-nucleotide polymorphisms, SNP) and multi-allelic polymorphisms (variable number of tandem repeats, VNTR and short tandem repeats, STR). This review describes the purpose of genetic markers in forensic investigation and their limitations. The STR loci are currently the most informative genetic markers for identity testing, but in cases without a suspect SNP can predict offender’s ancestry and phenotype traits such as skin, eyes and hair color. Nowadays, many countries worldwide have established forensic DNA databases based on autosomal short tandem repeats and other markers. In order for DNA profile database to be useful at a national or international level, it is essential to standardize genetic markers used in laboratories

Extramedullary involvement of lymph nodes in multiple myeloma

© 2017, University of Kragujevac, Faculty of Science. All rights reserved. Myeloma multiplex is a malignant disease of bone marrow plasma cells. It is usually confined to the bone marrow, but in rare cases, patients can develop extramedullary disease. The involvement of lymph nodes is rare and can be a diagnostic challenge. Here, we describe a 36-year-old male patient who presented with abdominal pain and discomfort initially. An abdominal ultrasound followed by computed tomography (CT) revealed retroperitoneal and mesenteric lymph node enlargement. Biopsies of the abdominal lymph node and infiltrated colon showed a plasma cell infiltrate positive for CD79α, CD38, CD138, kappa light chain and VEGF2. Multiple myeloma with extramedullary localization was diagnosed. After six cycles of chemotherapy consisting of doxorubicin, dexamethasone and thalidomide followed by autologous haematopoietic cell transplantation, the patient achieved complete remission. Specifically, a CT scan after therapy showed enlarged lymph nodes in the abdomen, but PET CT scans did not detect any metabolically active foci. Three years after the completion of therapy, the patient remains in remission. This case illustrates a rare presentation of extramedullary myeloma involving the abdominal lymph nodes, which could have been potentially mistaken for a lymphoid malignancy

Length of vegetation period as parameter of common oak (Quercus robur L.) phenological variability

Paper presents results for length of vegetation period as factor in common oak’s (Quercus robur L.) phenological variability. Trees in two common oaks populations on two locations in area of Belgrade were researched. Average length of vegetation period on site Ada Ciganlija is 166 days, while in location Bojčinska šuma is 179 days. Regarding that populations are in similar environmental and habitual conditions, differences in results should be treated as consequence of common oaks intra-specific variability and genetic structure of population. Significant differences in three consecutive years of observation are confirmed