Modulation of stress and immune response by Amblyomin-X results in tumor cell death in a horse melanoma model

We have investigated Amblyomin-X-treated horse melanomas to better understand its mode of action through transcriptome analysis and the in vivo model. Amblyomin-X is a Kunitz-type homologous protein that selectively leads to the death of tumor cells via ER stress and apoptosis, currently under investigation as a new drug candidate for cancer treatment. Melanomas are immunogenic tumors, and a better understanding of the immune responses is warranted. Equine melanomas are spontaneous and not so aggressive as human melanomas are, as this study shows that the in vivo treatment of encapsulated horse melanoma tumors led to a significant reduction in the tumor size or even the complete disappearance of the tumor mass through intratumoral injections of Amblyomin-X. Transcriptome analysis identified ER- and mitochondria-stress, modulation of the innate immune system, apoptosis, and possibly immunogenic cell death activation. Interactome analysis showed that Amblyomin-X potentially interacts with key elements found in transcriptomics. Taken together, Amblyomin-X modulated the tumor immune microenvironment in different ways, at least contributing to induce tumor cell death.

Modulation of stress and immune response by Amblyomin-X results in tumor cell death in a horse melanoma model

We have investigated Amblyomin-X-treated horse melanomas to better understand its mode of action through transcriptome analysis and the in vivo model. Amblyomin-X is a Kunitz-type homologous protein that selectively leads to the death of tumor cells via ER stress and apoptosis, currently under investigation as a new drug candidate for cancer treatment. Melanomas are immunogenic tumors, and a better understanding of the immune responses is warranted. Equine melanomas are spontaneous and not so aggressive as human melanomas are, as this study shows that the in vivo treatment of encapsulated horse melanoma tumors led to a significant reduction in the tumor size or even the complete disappearance of the tumor mass through intratumoral injections of Amblyomin-X. Transcriptome analysis identified ER- and mitochondria-stress, modulation of the innate immune system, apoptosis, and possibly immunogenic cell death activation. Interactome analysis showed that Amblyomin-X potentially interacts with key elements found in transcriptomics. Taken together, Amblyomin-X modulated the tumor immune microenvironment in different ways, at least contributing to induce tumor cell death.

Preclinical evaluation of Amblyomin-X, a Kunitz-type protease inhibitor with antitumor activity

Amblyomin-X, a Kunitz-type protease inhibitor, is a recombinant protein that selectively induces apoptosis in tumor cells and promotes tumor reduction in vivo in melanoma animal models. Furthermore, Amblyomin-X was able to drastically reduce lung metastasis in a mice orthotopic kidney tumor model. Due to its antitumor activity, Amblyomin-X potential to become a new drug is currently under investigation, therefore the aim of the present study was to perform preclinical assays to evaluate Amblyomin-X toxicity in healthy mice. Exploratory toxicity assays have shown that treatment with 512?mg/kg of Amblyomin-X lead to animal mortality, therefore two groups of treatment were evaluated in the present work: in the acute toxicity assay, animals were injected once with doses ranging from 4 to 256?mg/kg of Amblyomin-X, while in the subacute toxicity assay, animals were injected with 0.25, 0.57 and 1?mg/kg of Amblyomin-X daily, during 28 days. Following this treatment regimens, Amblyomin-X did not cause any mortality; moreover, toxicity signs were discrete, reversible and observed only at the higher doses, thus establishing a safety profile for administration in mice, which can be further used to determine the dose translation of this novel drug candidate for treatment in other species.

Preclinical evaluation of Amblyomin-X, a Kunitz-type protease inhibitor with antitumor activity

Amblyomin-X, a Kunitz-type protease inhibitor, is a recombinant protein that selectively induces apoptosis in tumor cells and promotes tumor reduction in vivo in melanoma animal models. Furthermore, Amblyomin-X was able to drastically reduce lung metastasis in a mice orthotopic kidney tumor model. Due to its antitumor activity, Amblyomin-X potential to become a new drug is currently under investigation, therefore the aim of the present study was to perform preclinical assays to evaluate Amblyomin-X toxicity in healthy mice. Exploratory toxicity assays have shown that treatment with 512?mg/kg of Amblyomin-X lead to animal mortality, therefore two groups of treatment were evaluated in the present work: in the acute toxicity assay, animals were injected once with doses ranging from 4 to 256?mg/kg of Amblyomin-X, while in the subacute toxicity assay, animals were injected with 0.25, 0.57 and 1?mg/kg of Amblyomin-X daily, during 28 days. Following this treatment regimens, Amblyomin-X did not cause any mortality; moreover, toxicity signs were discrete, reversible and observed only at the higher doses, thus establishing a safety profile for administration in mice, which can be further used to determine the dose translation of this novel drug candidate for treatment in other species.

Combination therapy of canine osteosarcoma with canine bone marrow stem cells, bone morphogenetic protein and carboplatin in an in vivo model

Osteosarcoma (OSA) is the most common malignant bone cancer in children and dogs. The ther-apeutic protocols adopted for dogs and humans are very similar, involving surgical options suchas amputation. Besides surgical options, radiotherapy and chemotherapy also are adopted. How-ever, hematologic, gastrointestinal and renal toxicity may occur because of chemotherapy treat-ments. Recent study clearly showed that mesenchymal stem cells (MSCs) combined withrecombinant human bone morphogenetic protein (rhBMP-2) may be associated with decreasesof the tumorigenic potential of canine OSA. The aim of this study was to analyse the efficacy ofchemotherapy with carboplatin and rhBMP-2 with MSCs in a canine OSA in vivo model. CanineOSA cells were implanted in mice Balb-c/nude with MSCs, rhBMP-2 and carboplatin. Flow cyto-metry and PCR for markers involved in tumour suppression pathways were analysed. Resultsshowed that the combination of MSCs and rhBMP-2 reduced tumour mass and infiltration ofneoplastic cells in tissues more efficiently than carboplatin alone. Thus it was demonstrated thatthe use of rhBMP-2 and MSCs, in combination with conventional antineoplastic, may be an effi-cient treatment strategy.

Quantitative evaluation of collagen and elastic fibers after intense pulsed light treatment of mouse skin

Background and Objective The aging of human skin includes intrinsic aging and photo-aging, which are characterized by a decrease in collagen and the deposition of abnormal elastic fibers. Intense pulsed light (IPL) sources are widely used in medicine to treat various cosmetic problems, including photo-damaged skin. Few studies have examined the microscopic changes produced by IPL. The objective of this study was to quantitatively evaluate the effects of IPL on collagen and elastic fibers in mice. Materials and Methods Forty female BALB/c mice were divided into four subgroups. Group 1 was the control group (n?=?10), and groups 2, 3, and 4 were treatment groups (n?=?10 in each group). Group 2 received one treatment, group 3 received two treatments, and group 4 received three treatments every 2 weeks. Skin tissue was obtained from irradiated areas 24 hours after the last treatment in each mouse. Collagen fibers were identified using the picrosirius red method. Elastic fibers were marked by Weigert-oxone stain. All samples were analyzed and quantified by a light microscope using analyzer system images. Results Group 4, which received three IPL treatments, showed significant quantitative increases in both collagen fibers (P<0.05) and elastic fibers (P<0.01). Collagen fibers demonstrated a better parallel distribution in relation to the epidermis. Conclusion IPL treatment significantly increased the number of collagen and elastic fibers within the dermis and improved the parallel distribution of collagen fibers in relation to the epidermis. These results were evident after three IPL treatments.

Chick embryo model for homing and host interactions of tissue engineering-purposed human dental stem cells

Human dental stem cells (hDSC) have a potential for regenerative therapies and could differentiate in vitro into many tissues, such as dentin, nerve, and vascular endothelium. Gallus gallus domesticus developing fertilized egg or chick embryo is an experimental model absent of xenografts rejection, largely employed in replacement of mammal species in scientific research and preclinical studies to evaluate angiogenesis and vasculogenesis, tissue differentiation, and embryonic development. This multiscale research deals with the homing and cell signaling effects of a standardized hDSC toward the receptor tissues of G. gallus domesticus in ovo. The hDSC were obtained from the explantation from third molars, characterized by cell cytometry, and employed without any further purification procedure. Four experimental groups were studied, according to the kind of cell tracing strategy, named: Control, mCherry-labeled hDSC, QTracker-labeled hDSC, and QTracker-exposed controls. The eggs were kept in an incubator temperature of 37.6 degrees C and humidity 86%, and the embryos were euthanized after 10 days of incubation. In vivo fluorescence and histological analysis were conducted. The fluorescence of the embryos inoculated with mCherry hDSC or the QTracker hDSC was associated with the bones and the beak tooth, and labeled cell islands could be localized in part of the samples. The inoculation of the QTracker probe resulted in proliferating bone tissue labeling. The hDSC inoculated groups presented cartilage plate hypertrophy and atypical morphology, meanwhile Control eggs were negative. The results demonstrated that hDSC can migrate to the cartilaginous tissues of the chick embryos, survive in this environment, implant, and interfere with the growth of developing bone.

Combination therapy of canine osteosarcoma with canine bone marrow stem cells, bone morphogenetic protein and carboplatin in an in vivo model

Osteosarcoma (OSA) is the most common malignant bone cancer in children and dogs. The ther-apeutic protocols adopted for dogs and humans are very similar, involving surgical options suchas amputation. Besides surgical options, radiotherapy and chemotherapy also are adopted. How-ever, hematologic, gastrointestinal and renal toxicity may occur because of chemotherapy treat-ments. Recent study clearly showed that mesenchymal stem cells (MSCs) combined withrecombinant human bone morphogenetic protein (rhBMP-2) may be associated with decreasesof the tumorigenic potential of canine OSA. The aim of this study was to analyse the efficacy ofchemotherapy with carboplatin and rhBMP-2 with MSCs in a canine OSA in vivo model. CanineOSA cells were implanted in mice Balb-c/nude with MSCs, rhBMP-2 and carboplatin. Flow cyto-metry and PCR for markers involved in tumour suppression pathways were analysed. Resultsshowed that the combination of MSCs and rhBMP-2 reduced tumour mass and infiltration ofneoplastic cells in tissues more efficiently than carboplatin alone. Thus it was demonstrated thatthe use of rhBMP-2 and MSCs, in combination with conventional antineoplastic, may be an effi-cient treatment strategy.

Quantitative evaluation of collagen and elastic fibers after intense pulsed light treatment of mouse skin

Background and Objective The aging of human skin includes intrinsic aging and photo-aging, which are characterized by a decrease in collagen and the deposition of abnormal elastic fibers. Intense pulsed light (IPL) sources are widely used in medicine to treat various cosmetic problems, including photo-damaged skin. Few studies have examined the microscopic changes produced by IPL. The objective of this study was to quantitatively evaluate the effects of IPL on collagen and elastic fibers in mice. Materials and Methods Forty female BALB/c mice were divided into four subgroups. Group 1 was the control group (n?=?10), and groups 2, 3, and 4 were treatment groups (n?=?10 in each group). Group 2 received one treatment, group 3 received two treatments, and group 4 received three treatments every 2 weeks. Skin tissue was obtained from irradiated areas 24 hours after the last treatment in each mouse. Collagen fibers were identified using the picrosirius red method. Elastic fibers were marked by Weigert-oxone stain. All samples were analyzed and quantified by a light microscope using analyzer system images. Results Group 4, which received three IPL treatments, showed significant quantitative increases in both collagen fibers (P<0.05) and elastic fibers (P<0.01). Collagen fibers demonstrated a better parallel distribution in relation to the epidermis. Conclusion IPL treatment significantly increased the number of collagen and elastic fibers within the dermis and improved the parallel distribution of collagen fibers in relation to the epidermis. These results were evident after three IPL treatments.

Chick embryo model for homing and host interactions of tissue engineering-purposed human dental stem cells

Human dental stem cells (hDSC) have a potential for regenerative therapies and could differentiate in vitro into many tissues, such as dentin, nerve, and vascular endothelium. Gallus gallus domesticus developing fertilized egg or chick embryo is an experimental model absent of xenografts rejection, largely employed in replacement of mammal species in scientific research and preclinical studies to evaluate angiogenesis and vasculogenesis, tissue differentiation, and embryonic development. This multiscale research deals with the homing and cell signaling effects of a standardized hDSC toward the receptor tissues of G. gallus domesticus in ovo. The hDSC were obtained from the explantation from third molars, characterized by cell cytometry, and employed without any further purification procedure. Four experimental groups were studied, according to the kind of cell tracing strategy, named: Control, mCherry-labeled hDSC, QTracker-labeled hDSC, and QTracker-exposed controls. The eggs were kept in an incubator temperature of 37.6 degrees C and humidity 86%, and the embryos were euthanized after 10 days of incubation. In vivo fluorescence and histological analysis were conducted. The fluorescence of the embryos inoculated with mCherry hDSC or the QTracker hDSC was associated with the bones and the beak tooth, and labeled cell islands could be localized in part of the samples. The inoculation of the QTracker probe resulted in proliferating bone tissue labeling. The hDSC inoculated groups presented cartilage plate hypertrophy and atypical morphology, meanwhile Control eggs were negative. The results demonstrated that hDSC can migrate to the cartilaginous tissues of the chick embryos, survive in this environment, implant, and interfere with the growth of developing bone.