21 research outputs found

    Human small cell lung cancer cell line (NCI-H146) tumor behaviour on the chicken embryo chorioallantoic membrane after the sodium valproate treatment

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    Small cell lung cancer (SCLC) is an aggressive form of cancer with 5-year survival rate and poor prognosis. Since patients' initial response to therapy is rapidly followed by a relapse with the drug-resistant disease, new therapies are required. The aim was to evaluate whether H146 cell line tumor was able to maintain the morphological pattern when grafted on the chicken embryo chorioallantoic membrane (CAM). Also, to evaluate the morphological changes in the CAM after grafting the tumor without or with treatment with sodium valproate (NaVP). The cell culture of the commercial NCIH146 cell line was used for the formation of the tumor. The tumor of 1x106 cells and I type rat tail collagen was dropped onto the absordable sponge and instantly grafted on CAM. After 5 days of incubation chicken embryos were sacrificed and CAMs were cut out. Standard H&E staining and immunohistochemistry (CD56) were performed. The histomorphometrical analysis of non-treated (n=5), 4 mM NaVP-treated (n=5) and 8 mM (n=5) of NaVP-treated groups was performed. The thickness of the CAM was measured in all the investigated groups in the areas under the onplant and in the neighbouring sites. H146 cells are able to maintain morphological appearance on the CAM and retained the expression of specific profile for CD56. H146 cell tumor increased the thickness of the CAM under the tumor in the non-treated group (509.2±184μm). After treatment with 4 mM and 8 mM of NaVP CAM thickness decreased (362±232.6 μm and 147.8±98.4 μm respectively; p<0.0001). The effect of NaVP on CAM increased with increasing solution concentrations of 4 mM and 8 mM. CAM thickness in the non-treated group neighbouring to the tumor site was 139.1±138.4 μm, in 4 mM NaVP-treated group – 104.9±81 μm and in 8 mM NaVP-treated group CAM thickness reached only 70.8±51.5 μm. CAM thickness in the non-treated group significantly differs compared to 4 mM and 8 mM of NaVP – the treated groups (p<0.05, p<0.0001). Difference between both NaVP-treated groups is also significant (p<0.05). CAM is a suitable model for in vivo analysis of human H146 cell line formed tumor. Cells grafted on the CAM preserved their morphology and expressed characteristic immunohistochemical markers. CAM mesenchyme proliferation was suppressed under the influence of NaVP in concentration–dependent manner

    Evaluation of the chicken embryo chorioallantoic membrane model for laryngeal tumor transplantation

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    The laryngeal squamous cell carcinoma is the second common malignant tumor of the respiratory tract and together with recurrent respiratory papillomas represents the most common tumors of the larynx. Many experimental models are used to study the morphology of malignant tumors. The chicken chorioallantoic membrane (CAM) model is one of them. The CAM has all the nutrition needed for the piece of the transplanted tumor to survive. The aim of this study was to investigate whether the laryngeal papilloma and the laryngeal squamous cell carcinoma tissues transplanted on the chick CAM survive with their main histological features, and to determine the morphological changes of the CAM with different transplants. For the preparation of the CAM, fertilized hen eggs were put into an incubator for 3 days. Then the windows in the shell were opened. The fresh samples of tumors were transplanted on the CAM on the 7th day of incubation. After 3 days after transplantation the CAM with onplants were excised and fixed in the 10% formalin solution. Morphological changes in the control CAM and in the CAM with tumor onplants were observed using the digital camera on the OLYMPUS microscope. The results showed that the CAM with the laryngeal squamous cell carcinoma onplant was distinctly thicker than that of the control group and than the CAM with the papilloma onplant; the chorionic epithelium was thickened and appeared stratified of up to 5–6 layers and in some locations squamous keratinized; the mesenchymal cells were densely arranged under the tumor transplants. We observed that morphological changes in the thickness of the CAM and the chorionic epithelium were more obvious in the CAM under the carcinoma transplants. After 72 hours of the tumor tissue transfer onto the membrane, the tumor cells retained their vitality and also their influence on the CAM tissues could be observed

    Mozaikinė autologinė kremzlės ir kaulo transplantacija gydant kelio sąnario kremzlės pažeidimus

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    Rimtautas Gudas1, Romas Jonas Kalesinskas1, Giedrius Bernotavičius1, Eglė Monastyreckienė2, Angelija Valančiūtė3, Darius Pranys41 Kauno medicinos universiteto klinikųOrtopedijos ir traumatologijos klinikaEivenių g. 2, LT-50009 Kaunasel paštas: [email protected] Kauno medicinos universiteto klinikųRadiologijos klinika3 Kauno medicinos universitetoEmbriologijos ir histologijos katedra4 Kauno medicinos universitetoPatologinės anatomijos klinika Tikslas Mūsų prospektyvaus klinikinio tyrimo tikslas – įvertinti mozaikinės autologinės transplantacijos rezultatus gydant kelio sąnario kremzlės pažeidimus. Ligoniai ir metodai 1998–2002 metais KMU Ortopedijos ir traumatologijos klinikoje atliktos 87 autologinės mozaikinės transplantacijos operacijos esant kelio sąnario kremzlės ir kaulo pažeidimams. Pacientų amžiaus vidurkis operacijų metu buvo 24,74 ± 7,20 metų (14–40 metų). Simptomai vidutiniškai truko 21,32 ± 5,57 mėnesio, o pacientai įvertinti praėjus vidutiniškai 24,4 mėnesio (nuo 12 iki 60 mėnesių) po operacijos. Rezultatai apibendrinti naudojant ICRS (International Cartilage Repair Society) anketą, 13 (15%) atvejų – remiantis kartotinių artroskopijų metu atliktu makroskopiniu įvertinimu, 9 (11%) atvejais – histologiniu tyrimu, 57 (67%) atvejais – BMR ir visais atvejais – rentgeno tyrimais. Visus anketinius įvertinimus atliko nepriklausomas gydytojas ortopedas prieš operacijas ir po operacijų praėjus dvylikai, dvidešimt keturiems, trisdešimt šešiems, keturiasdešimt aštuoniems ir šešiasdešimčiai mėnesių. Nepriklausomi nuo tyrimo gydytojai, radiologas ir patologas, atliko visus BMR ir histomorfologinius įvertinimus. Rezultatai Praėjus vidutiniškai 24,4 mėnesio po mozaikinės transplantacijos, ICRS anketos būdu nustatytas klinikinis rezultatų pagerėjimas (p < 0,05). Funkcinis ir objektyvus įvertinimas pagal ICRS anketą buvo toks: 93% atvejų gauti geri ir labai geri operacinio gydymo rezultatai, kitais atvejais (7%) gydymo rezultatai buvo patenkinami. Kartotinių artroskopijų metu (remiantis ICRS protokolu), po operacijų praėjus vidutiniškai 12,4 mėnesio, 11 (84%) iš 13 atvejų buvo nustatytas geras ir labai geras sąnario kremzlės makroskopinis atsitaisymas. Sąnario kremzlės biopsijos ir histologinis tyrimas buvo atliktas 11% pacientų, ir daugumos jų (remiantis ICRS protokolu) sąnario kremzlės ir pokremzlinio kaulo atitaisymo koeficientas buvo geras. BMR tyrimas parodė, kad 94% pacientų sąnario paviršius atkurtas gerai arba labai gerai. Išvados Mūsų tyrimas parodė, kad po mozaikinės autologinės transplantacijos praėjus vidutiniškai 24,4 mėnesio (nuo 12 iki 60 mėn.) fiziškai aktyvių pacientų, kuriems buvo pažeista kelio sąnario kremzlė, klinikinė ir funkcinė būklė labai pagerėjo. Dauguma pacientų po kremzlės mozaikinės autologinės transplantacijos atgauna prieš pažeidimą buvusį fizinio aktyvumo lygį. Histomorfologinis persodintų transplantatų tyrimas parodė, kad visais tirtais atvejais išsilaikė hialininės kremzlės struktūra ir fibroelastinio audinio intarpai tarp transplantatų. Reikšminiai žodžiai: sąnario kremzlės pažeidimai, mozaikinė transplantacija Mosaic-like autologous osteochondral transplantation for the treatment of knee joint articular cartilage injuries Rimtautas Gudas1, Romas Jonas Kalesinskas1, Giedrius Bernotavičius1, Eglė Monastyreckienė2, Angelija Valančiūtė3, Darius Pranys4 Objective The purpose of this prospective clinical study was to evaluate the outcomes of mosaic type autologous osteochondral transplantation procedure for the treatment of the articular cartilage defects of the knee joint. Patients and methods Between 1998 and 2002, a total of 85 patients underwent an osteochondral autologous transplantation (mosaicplasty) procedure for osteochondral or chondral knee joint injury. The patients were evaluated using the ICRS score, arthroscopically, histologically, with MRI and X-ray examinations. The mean duration of symptoms was 21.32 ± 5.57 months and the mean follow-up was 24.4 months (range, 12 to 60 months). The mean age of the patients during the surgery was 24.74 ± 7.20 years (range, 14 to 40 years). An independent observer performed a follow-up examination after six, twelve, twenty-four, thirty-six and forty-eight months. In 13 (15%) of 85 cases 12.4 months postoperatively, arthroscopy with biopsy for histological evaluation was carried out. A radiologist and a pathologist, both blinded to each patient’s treatment, did the radiological and histological evaluations. Results After 24.4 months all the patients showed a significant clinical improvement (p < 0.05). The cartilage Repair Society (ICRS) score, functional and objective assessment revealed 93% to have excellent or good results after mosaicplasty; 7% were fair 24.4 months (range, 12–60 months) after the operations. The ICRS evaluation showed a significant improvement after the mosaicplasty procedure 24.4 months following operations (p = 0.005). No serious complications were reported. The ICRS for macroscopic evaluation during arthroscopy 12.4 months after mosaicplasty demonstrated excellent or good repair in 11 (84%) of 13 cases. Biopsy specimens were obtained from 9 (69%) of 13 patients, and histological evaluation of repair showed good scores (according to ICRS) for most samples after mosaicplasty. A MRI evaluation demonstrated excellent or good repairs in 94% after mosaicplasty. Conclusions On an average after 24.4 months (range, 12 to 60 months) of follow-up, our clinical study has shown a significant improvement of the clinical status of the physically active patients after the mosaic type autologous osteochondral transplantation for the repair of articular cartilage defects in the knee. Histologically, the osteochondral cylinder transplants retained hyaline cartilage. Keywords: articular cartilage injury, osteochondral mosaic-like transplantatio

    Collagen network changes in basilar artery in aging

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    Objective. The aim of this study was to examine and evaluate morphometrically age-related changes in collagen network in the tunica media of human basilar artery. Materials and methods. Histological analysis of the basilar arteries obtained from 89 individuals, aged 20 to 85 years, was performed. The study has been carried out by standard histological technique; histological samples were stained with picrosirius red. Detailed quantitative analysis of collagen bundle network in the tunica media of basilar artery was performed. Results. We analyzed collagen network area, perimeter and number of collagen bundles in the tunica media of the basilar artery. After investigation of age-related changes in collagen network area in men and women, we determined that in both genders collagen network area in the tunica media of the basilar artery increased with age. Analyzing the perimeter and number of collagen bundles in both genders by different age groups, it was found that they decreased with age. We found a statistically significant correlation between all the measured parameters and the age. Conclusions. The area of collagen bundles increased, the number and perimeter of collagen bundles decreased with age in both genders in the tunica media of human basilar artery. While aging, the structure of collagen bundle network became less branchy, collagen fibers merged, their cross section area enlarged

    Sodium Valproate Inhibits Small Cell Lung Cancer Tumor Growth on the Chicken Embryo Chorioallantoic Membrane and Reduces the p53 and EZH2 Expression

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    The study aims to test the effect of different sodium valproate (NaVP) doses on small cell lung cancer NCI-H146 cells tumor in chicken embryo chorioallantoic membrane (CAM) model. Xenografts were investigated in the following groups: nontreated control and 5 groups treated with different NaVP doses (2, 3, 4, 6, and 8 mmol/L). Invasion of tumors into CAM in the nontreated group reached 76%. Tumors treated with 8 mmol/L NaVP doses significantly differed in tumor invasion frequency from the control and those treated with 2 mmol/L ( P < .01). The calculated probability of 50% tumor noninvasion into CAM was when tumors were treated with 4 mmol/L of NaVP. Number of p53-positive cells in tumors was significantly reduced when treated with NaVP doses from 3 to 8 mmol/L as compared with control; number of EZH2-positive cells in control significantly differed from all NaVP-treated groups. No differences in p53- and EZH2-positive cell numbers were found among 4, 6, and 8 mmol/L NaVP-treated groups. Invaded tumors had an increased N-cadherin and reduced E-cadherin expression. The results indicate the increasing NaVP dose to be able to inhibit tumors progression. Expression of p53 and EZH2 may be promising target markers of therapeutic efficacy evaluation

    Different Effects of Valproic Acid on SLC12A2, SLC12A5 and SLC5A8 Gene Expression in Pediatric Glioblastoma Cells as an Approach to Personalised Therapy

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    Valproic acid (VPA) is a histone deacetylase inhibitor with sex-specific immunomodulatory and anticancer effects. This study aimed to investigate the effect of 0.5 and 0.75 mM VPA on NKCC1 (SLC12A2), KCC2 (SLC12A5) and SLC5A8 (SLC5A8) co-transporter gene expressions in pediatric PBT24 (boy&rsquo;s) and SF8628 (girl&rsquo;s) glioblastoma cells. The SLC12A2, SLC12A5 and SLC5A8 RNA expressions were determined by the RT-PCR method. The SLC12A2 and SLC5A8 expressions did not differ between the PBT24 and SF8628 controls. The SLC12A5 expression in the PBT24 control was significantly higher than in the SF8628 control. VPA treatment significantly increased the expression of SLC12A2 in PBT24 but did not affect SF8628 cells. VPA increased the SLC12A5 expression in PBT24 and SF8628 cells. The SLC12A5 expression of the PBT24-treated cells was significantly higher than in corresponding SF8628 groups. Both VPA doses increased the SLC5A8 expression in PBT24 and SF8628 cells, but the expression was significantly higher in the PBT24-treated, compared to the respective SF8628 groups. The SLC5A8 expression in PBT24-treated cells was 10-fold higher than in SF8628 cells. The distinct effects of VPA on the expression of SLC12A2, SLC12A5 and SLC5A8 in PBT24 and SF8628 glioblastoma cells suggest differences in tumor cell biology that may be gender-related

    Osteochondral transplantation for the treatment of femoral condyle defects

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    Between 1998 and 2001 thirty-five patients underwent osteochondral transplantation (mosaicplasty) and 35 patients (controls) \u96 microfracture procedure for osteochondral or chondral knee joint pathology treatment. Average age of patients was 24.74±7.20 years. Patients were evaluated through International Cartilage Repair Society (ICRS) and modified Hospital for special Surgery (HSS) scales, arthroscopically, histologically, with MRI and x-rays. Modified HSS and ICRS evaluation showed statistically significantly better results in the mosaicplasty group 12 months post operation (p=0.005). Last follow-up showed deterioration in microfracture group (p=0.0005)

    Model of human recurrent respiratory papilloma on chicken embryo chorioallantoic membrane for tumor angiogenesis research

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    We aimed to develop a chick embryo chorioallantoic membrane (CAM) model of recurrent respiratory papilloma (RPP) and to evaluate its morphological and morphometric characteristics, together with angiogenic features. Fresh RRP tissue samples obtained from 13 patients were implanted in 174 chick embryo CAMs. Morphological, morphometric, and angiogenic changes in the CAM and chorionic epithelium were evaluated up until 7 days after the implantation. Immunohistochemical analysis (34βE12, Ki-67, MMP-9, PCNA, and Sambucus nigra staining) was performed to detect cytokeratins and endothelial cells and to evaluate proliferative capacity of the RRP before and after implantation on the CAM. The implanted RRP tissue samples survived on CAM in 73% of cases while retaining their essential morphologic characteristics and proliferative capacity of the original tumor. Implants induced thickening of both the CAM (241-560%, p=0.001) and the chorionic epithelium (107-151%, p=0.001), while the number of blood vessels (37-85%, p=0.001) in the CAM increased. The results of the present study confirmed that chick embryo CAM is a relevant host for serving as a medium for RRP fresh tissue implantation. The CAM assay demonstrated the specific RRP tumor growth pattern after implantation and provided the first morphological and morphometric characterization of the RRP CAM model that opens new horizons in studying this disease

    The Importance of gender-related anticancer research on mitochondrial regulator sodium dichloroacetate in preclinical studies In Vivo

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    Sodium dichloroacetate (DCA) is an investigational medicinal product which has a potential anticancer preparation as a metabolic regulator in cancer cells’ mitochondria. Inhibition of pyruvate dehydrogenase kinases by DCA keeps the pyruvate dehydrogenase complex in the active form, resulting in decreased lactic acid in the tumor microenvironment. This literature review displays the preclinical research data on DCA’s effects on the cell pyruvate dehydrogenase deficiency, pyruvate mitochondrial oxidative phosphorylation, reactive oxygen species generation, and the Na+–K+–2Cl− cotransporter expression regulation in relation to gender. It presents DCA pharmacokinetics and the hepatocarcinogenic effect, and the safety data covers the DCA monotherapy efficacy for various human cancer xenografts in vivo in male and female animals. Preclinical cancer researchers report the synergistic effects of DCA combined with different drugs on cancer by reversing resistance to chemotherapy and promoting cell apoptosis. Researchers note that female and male animals differ in the mechanisms of cancerogenesis but often ignore studying DCA’s effects in relation to gender. Preclinical gender-related differences in DCA pharmacology, pharmacological mechanisms, and the elucidation of treatment efficacy in gonad hormone dependency could be relevant for individualized therapy approaches so that gender-related differences in treatment response and safety can be proposed

    Tumors derived from lung cancer cells respond differently to treatment with sodium valproate (a HDAC inhibitor) in a chicken embryo chorioallantoic membrane model

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    Lung cancer is the most frequent cause of cancer death. Some human lung malignant tumors have a combined small-cell lung cancer (SCLC) and nonsmall-cell lung cancer (NSCLC) histology, with tumor cell phenotype changing during tumor progression. Valproic acid is used as an anti-seizure medication to treat migraine, and bipolar mood disorders. Recently, its efficacy as an adjuvant therapy was shown in cancer due to its histone deacetylase (HDAC) inhibitory property. HDACs are upregulated in lung tumors, and HDAC inhibitors, including valproic acid, inhibit endothelial cell proliferation in vitro and in vivo and have antiproliferative and antimigratory properties. We tested valproic acid for possible antiangiogenic and antimigratory effects on experimental lung tumors grafted onto the chicken embryo chorioallantoic membrane (CAM). Tumors were formed from two NSCLC cell lines and a single SCLC cell line. To investigate tumor and CAM interactions, in vivo biomicroscopy, visualization of blood vessels with injected fluorescent dextran, histological, immunohistochemical and histomorphometric methods were applied. Our results showed that a sodium valproate (NaVP) treatment-induced a dose-dependent decrease of experimental tumor invasion into the CAM mesenchyme and a reduction in angiogenesis. Both the invasion and the angiogenic response were dependent on the type of cell line used: invasion and angiogenesis of tumors derived from A549 and NCI-H146 cell lines responded to increasing doses of NaVP from 4 to 8 mM, whereas Sk_Lu_1 cells response were antimigratory and antiangiogenic when NaVP was used up to 6 mM. When 8mM NaVP was used, stimulated invasion and angiogenesis in tumors from Sk_Lu_1 cells were observed
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