scholarly journals 5-ALA-mediated fluorescence of musculoskeletal tumors in a chick chorio-allantoic membrane model: preclinical in vivo qualification analysis as a fluorescence-guided surgery agent in Orthopedic Oncology

2022 ◽  
Vol 17 (1) ◽  
Author(s):  
Wiebke K. Guder ◽  
Wolfgang Hartmann ◽  
Clarissa Buhles ◽  
Maike Burdack ◽  
Maike Busch ◽  
...  
Keyword(s):  
Cell Culture ◽  
Primary Cell Culture ◽  
Aminolevulinic Acid ◽  
Primary Cell ◽  
Guided Surgery ◽  
Musculoskeletal Tumors ◽  
Fluorescence Guided Surgery ◽  
Orthopedic Oncology ◽  
Macroscopic Tumor

Abstract Background Fluorescence-guided surgery (FGS) with 5-aminolevulinic acid (5-ALA) and other contrast agents has shown its efficacy in improving resection margins, local recurrence and survival rates in several medical disciplines. It is the objective of this study to analyze the engraftment rate of musculoskeletal tumor specimens on the chick chorio-allantoic membrane (CAM), the rate of tumor fluorescence (PDD), and the effects of photodynamic therapy (PDT) after exposure of tumors to 5-ALA in an in vivo environment. Methods A total of 486 CAMs were inoculated with macroscopic tumor grafts (n = 26; n = 478 eggs) and primary cell culture suspensions (n = 2; n = 8 eggs) from 26 patients on day 10 of egg development. On day 16, 2 mg/200 µl 5-ALA were topically applied per egg. After 4 h of incubation, Protoporphyrin IX was excited using blue light (420 ± 10 nm). Tumor fluorescence (PDD) was photo-documented. A subgroup of specimens was additionally exposed to red light (635 nm ± 10 nm; PDT). After the termination of the experiment, CAM-grown tumors were histopathologically analyzed. Results Benign and borderline tumors (chondroblastoma, giant cell tumor of bone and atypical chondrogenic tumor) presented with high rates of detectable fluorescence. Comparable results were found for chondrosarcoma, osteosarcoma and Ewing’s sarcoma among bone and dedifferentiated liposarcoma, myxofibrosarcoma and undifferentiated pleomorphic sarcoma among soft tissue sarcomas. Overall, tumor fluorescence was negative for 20.2%, single-positive (+) for 46.9% and double-positive (++) for 32.9% of macroscopic xenografts, and negative in 20% and (+) in 80% of primary cell culture tumors. Macroscopic tumor xenografts (n = 478) were identified as viable in 14.8%, partially viable in 2.9% and partially to completely regressive in 45.2%. All (n = 8) tumors grown from primary cell culture were viable. After PDT, tumor samples were found viable in 5.5%, partially viable in 5.5% and partially to completely regressive in 68%. Egg survival increased with decreasing PDT doses. Conclusions The CAM model proves to be a suitable in vivo model for the investigation of short-term observation questions in musculoskeletal tumors. The findings of this study warrant further investigation of PDT effects on musculoskeletal tumors and a possible incorporation of 5-ALA FGS in clinical Orthopedic Oncology care.

Rapid melanization of bomirski amelanotic melanoma cells in cell culture

1983 ◽  
Vol 3 (2) ◽  
pp. 189-194 ◽  
Author(s):  
A. Słominski
Keyword(s):  
Cell Culture ◽  
Primary Cell Culture ◽  
Growth Conditions ◽  
Melanoma Cells ◽  
Amelanotic Melanoma ◽  
Primary Cell ◽  
In Vitro Growth

Transfer of Bomirski amelanotic melanoma ceils from in vivo to in vitro growth conditions results in occurrence of rapid melanization in their cytoplasm. The melanized ceils from primary cell culture initiate tumours in hamsters, which do not contain traces of melanin and resemble typical amelanotic melanoma.

scholarly journals Do alterations in gene expressions influence tumorigenesis in the transmissible venereal tumor in dogs?

2020 ◽  
Vol 50 (11) ◽  
Author(s):  
Haline Ballestero Fêo ◽  
Luis Mauricio Montoya Flórez ◽  
Ricardo Seiti Yamatogi ◽  
Anderson do Prado Duzanski ◽  
João Pessoa Araújo Junior ◽  
...  
Keyword(s):  
Cell Culture ◽  
Tumor Suppressor Genes ◽  
Primary Cell Culture ◽  
Gene Mutations ◽  
Tp53 Gene ◽  
Primary Cell ◽  
Loss Of Function ◽  
Gene Expressions

ABSTRACT: Canine transmissible venereal tumor (CTVT) is a transmissible neoplasm, which spreads naturally between dogs through the halogenic transfer of tumor cells, mainly during coitus. It is the oldest known tumoral lineage in nature and reports on gene mutations have been extended. Also, this tumor shares several genetic mutations with some cancers in humans, among them lung carcinomas, melanoma, prostate, breast, among other cancers. Thus, expression of tumor suppressor genes such as TP53, P21, and apoptosis-related genes such as BAX, BCL-2, and BCL-xL, both in vivo and in vitro (primary cell culture) were quantified. In the present study, the comparison of gene expression, the TP53 gene, in most cases, was shown to be high in the majority of tissues (65%) and primary cell culture (100%), while BCL-2, BCL-xL, and BAX presented variation among the animals analyzed. Moreover, in these situations, the results suggested that the apoptotic regulation of these genes did not occur for TP53. The P21 gene was shown to be mostly normal (70%); although, absence (6%) and underexpressions (24%) were also observed. Statistical analysis of the BCL-xL gene demonstrated significant differences between the tissues of the animals when compared to the cell cultures; however, to the other genes, no statistical difference was observed between the groups. Preliminarily, the results suggested the presence of alterations in the gene expressions of the TP53, P21, BAX, BCL-2 and BCL-xL leading to loss of function in these genes, which affect the tumorigenesis of CTVT.

scholarly journals Expression of Selected Connexin and Aquaporin Genes and Real-Time Proliferation of Porcine Endometrial Luminal Epithelial Cells in Primary Culture Model

2020 ◽  
Vol 2020 ◽  
pp. 1-15 ◽  
Author(s):  
Katarzyna Wojtanowicz-Markiewicz ◽  
Magdalena Kulus ◽  
Sandra Knap ◽  
Ievgenia Kocherova ◽  
Maurycy Jankowski ◽  
...  
Keyword(s):  
Cell Culture ◽  
Epithelial Cells ◽  
Primary Cell Culture ◽  
Water Movement ◽  
Inorganic Ions ◽  
Primary Cell ◽  
Lag Phase ◽  
Luminal Epithelial Cells

Luminal epithelial cells are the first embryonic-maternal contact site undergoing very specific changes associated with reproductive processes. Cells prepare for embryo development by increasing their volume, with the help of aquaporins that provide a transcellular path of rapid water movement during the secretion and absorption of fluids, as well as connexins enabling the flow of inorganic ions and small molecules. In this work, we have examined how AQPs and Cx’s behave in luminal epithelium primary cell culture. Cells obtained from porcine specimen during slaughter were primarily in vitro cultured for 7 days. Their proliferation patterns were then analyzed using RTCA, with the expression of genes of interest evaluated with the use of immunofluorescence and RT-qPCR. The results of these changes of gene of interest expression were analyzed on each of the seven days of the porcine luminal primary cell culture. Our study showed that the significant changes were noted in the case of Cx43, whose level of protein expression and distribution increases after 120 hours of culture, when the cells enter the lag phase, and maintains an upward trend until the end of the culture. We noted an increase in AQP4, AQP7, AQP8, and AQP11 levels throughout the entire culture period, while the largest differences in expression were found in AQP3, AQP4, and AQP10. The obtained results could become a point of reference for further in vivo and clinical research. Experiments conducted with these proteins showed that they influence the endometrial fluid content during the oestrous cycle and participate in the process of angiogenesis, which intensifies during endometrial development.

In Vivo-like model of glial tumors: Creation of primary cell lines.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e14515-e14515
Author(s):  
Sofia V. Timofeeva ◽  
Oleg I. Kit ◽  
Anastasia O. Sitkovskaya ◽  
Irina V. Mezhevova ◽  
Svetlana Yu. Filippova ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Lines ◽  
Primary Cell Culture ◽  
Tumor Tissue ◽  
Primary Cell ◽  
Glial Tumor ◽  
Ki 67 ◽  
Glial Tumors

e14515 Background: The choice of cell source for 3D bioprinting of in vivo-like models of glial tumors is crucial and must take into account the ability to proliferation and stable metabolism. Oral administration of 5-aminolevulinic acid (5-ALA) in patients prior to surgery increases the fluorescent contrast between tumor and surrounding tissue, but the effect of contrast agents on cells in vitro is unknown. The aim of the study was obtaining viable glial tumor tissues using 5-ALA, as well as the development of a stable primary cell culture for 3D bioprinting. Methods: Tumor tissue was obtained from patients with glioblastoma during surgery under visual control using the Opmi Pentero Blue E400 microscope and 5-ALA. Material was disaggregated on a BD Machine using Medicons 50 μm (BD). Glioblastoma cells were cultured in DMEM/F12 medium with L-glutamine (Gibco) containing 10% fetal bovine serum (Biolot, Russia), 1% non-essential amino acids (NEAA, Sigma-Aldrich) and 0.5% penicillin-streptomycin (Biolot) at 37C. Glial cell lines were characterized immunohistochemically using antibodies to the glial fibrillary acidic protein (GFAP) and proliferation index (Ki-67). Microsatellite analysis was performed using three dinucleotide repeat markers D2S123, D17S250, D5S346 and five mononucleotide loci BAT25, BAT26, NR21, NR24 and NR27. Results: The positive expression of GFAP on the cell processes of the star-like shape was clearly visualized, indicating a morphological feature of glial tumors. The Ki-67 labeling index was 70%. Changes were observed at the D17S250 locus (148-148/148-152) for the glial tumor primary cells after the sixth passage. Microsatellite instability was not observed in the primary cell culture. Conclusions: The accumulation of porphyrins from 5-ALA in glial tumor cells does not prevent the in vitro creation of a cell culture from tumor tissue. Microsatellite analysis showed that the obtained glioblastoma cell lines remain stable for at least 10 passages. Material obtained during resection using 5-ALA is a reliable source of stable glial tumor cell lines.

Preclinical evidence of ET-743 as a potential chemotherapy option for the treatment of biliary carcinoma.

2013 ◽  
Vol 31 (4_suppl) ◽  
pp. 193-193
Author(s):  
Francesco Leone ◽  
Caterina Peraldo-Neia ◽  
Giuliana Cavalloni ◽  
Marco Soster ◽  
Loretta Gammaitoni ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Culture ◽  
Dna Damage ◽  
Clinical Trials ◽  
Cell Lines ◽  
Primary Cell Culture ◽  
Primary Cell ◽  
Term Survival ◽  
Anti Tumor Activity

193 Background: The standard chemotherapy for unresectablebiliary tract carcinoma (BTC) is based on gemcitabine and platinum compounds. However, these combinations have not been shown to be effective in improving long-term survival. Thus, there is a real need to find new strategies that would impact in a significant way on clinical outcome. Ecteinascidin-743 (ET-743), a compound isolated from the marine tunicate Ecteinascidia turbinata. ET-743, is approved for the treatment of ovarian cancer and soft tissue sarcoma. Phase II and III clinical trials are ongoing for the treatment of different solid tumors. No preclinical data are available about the efficacy of ET-743 in BTC. In a phase I study, one patient received ET-743 plus capecitabine and experienced a long lasting complete metabolic response. Here, we investigated the antitumor activity of ET-743 in preclinical BTC models. Methods: Four BTC cell lines TFK1, EGI-1, HuH28 and TGBC1 were used to evaluate the effect of ET-743 on proliferation, cell cycle, apoptosis and on the activation of DNA damage proteins. The effect on proliferation was also investigated on a primary cell culture of a gallbladder carcinoma (GBC) resistant to gemcitabine and oxaliplatin. On the same cells, the inhibition of VEGF secretion mediated by ET-743 was analyzed by ELISA. The anti-tumor activity of ET-743 was tested on EGI-1 xenografts in NOD/SCID mice. Results: In vitro, ET-743 is able to markedly reduce cell proliferation of BTC cell lines through cell cycle blockage on G0/G1 phase and to inhibit the growth of primary cell culture derived from GBC patient. Moreover, ET-743 promotes apoptosis by caspase 3 activation, activates proteins involved in DNA damage and reduces VEGF secretion. In the in vivo model, ET-743 is able to slow tumor growth in BTC xenograft. The mechanism of anti-tumor activity involves DNA damage, the induction of hypoxia transcription factor-1, and angiogenesis inhibition. ET-743 has no significant effect on apoptosis in vivo. Conclusions: These data suggest that ET-743 could represent an alternative chemotherapy for BTC treatment and encourage the development of clinical trials of ET-743 in BTC patients.

scholarly journals Reverse Transport of Cholesterol Is the Reason for Resistance to Development of Atherosclerosis in Prague Hereditary Hypercholesterolemic (PHHC) Rat

2014 ◽  
pp. 591-596
Author(s):  
M. SCHMIEDTOVA ◽  
M. HECZKOVA ◽  
J. KOVAR ◽  
I. KRALOVA LESNA ◽  
R. POLEDNE
Keyword(s):  
Cell Culture ◽  
Adipose Tissue ◽  
Cholesterol Efflux ◽  
Wistar Rats ◽  
Primary Cell Culture ◽  
Primary Cell ◽  
Reverse Transport ◽  
Cholesterol Levels ◽  
Muscle Tissues

The Prague Hereditary Hypercholesterolemic (PHHC) rat is a model of hypercholesterolemia. In previous experiments, it was found to be completely resistant to the development of atherosclerosis. It was assumed that the reverse transport of cholesterol (RCT) might be the reason for this resistance. In this study, RCT was measured in vivo by cholesterol efflux from macrophages to plasma, using previously established methods for RCT in mice (Rader 2003), optimized for measurements in rats. Primary cell culture of macrophages was labeled with 14C-cholesterol and then injected intraperitoneally into rats. Plasma and feces were collected at 24 and 48 h. The plasma 14C-cholesterol levels at both 24 and 48 h were significantly higher in male PHHC rats compared to control Wistar rats. The PHHC rats excreted less 14C-cholesterol in feces in 24 and 48 h compared to Wistar rats. The largest pool of 14C-cholesterol was found in the adipose tissue of PHHC rats and in contrast lower levels of 14C-cholesterol were measured in the liver and muscle tissues of PHHC rats compared with Wistar rats. Increasing release of 14C-cholesterol efflux from macrophages demonstrates accelerated RTC and leads to prevention of atherogenesis in PHHC rats.

Optical imaging probes and their potential contribution to radiotracer development

2016 ◽  
Vol 55 (02) ◽  
pp. 51-62 ◽  
Author(s):  
S. Hermann ◽  
M. Schäfers ◽  
C. Höltke ◽  
A. Faust
Keyword(s):  
Optical Imaging ◽  
Fluorescent Dyes ◽  
Great Influence ◽  
Potential Contribution ◽  
Preclinical Evaluation ◽  
Guided Surgery ◽  
Fluorescence Guided Surgery ◽  
Imaging Probes ◽  
Unspecific Binding

SummaryOptical imaging has long been considered a method for histological or microscopic investigations. Over the last 15 years, however, this method was applied for preclinical molecular imaging and, just recently, was also able to show its principal potential for clinical applications (e.g. fluorescence-guided surgery). Reviewing the development and preclinical evaluation of new fluorescent dyes and target-specific dye conjugates, these often show characteristic patterns of their routes of excretion and biodistribution, which could also be interesting for the development and optimization of radiopharmaceuticals. Especially ionic charges show a great influence on biodistribution and netcharge and charge-distribution on a conjugate often determines unspecific binding or background signals in liver, kidney or intestine, and other organs.Learning from fluorescent probe behaviour in vivo and translating this knowledge to radio-pharmaceuticals might be useful to further optimize emerging and existing radiopharmaceuticals with respect to their biodistribution and thereby availability for binding to their targets.

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