Indoleamine-2,3 dioxygenase: a fate-changer of the tumor microenvironment.

Indoleamine-2,3 dioxygenase is a rate-limiting enzyme in the tryptophan catabolism in kynurenine pathways that has an immunosuppressive effect and supports cancer cells to evade the immune system in different cancer types. Diverse cytokines and pathways upregulate the production of indoleamine-2,3 dioxygenase enzymes in the tumor microenvironment and cause more production and activity of this enzyme. Ultimately, this situation results in anti-tumor immune suppression which is in favor of tumor growth. Several inhibitors such as 1-methyl-tryptophan have been introduced for indoleamine-2,3 dioxygenase enzyme and some of them are widely utilized in pre-clinical and clinical trials. Importantly at the molecular level, indoleamine-2,3 dioxygenase is positioned in a series of intricate signaling and molecular networks. Here, the main objective is to provide a focused view of indoleamine-2,3 dioxygenase enhancer pathways and propose further studies to cover the gap in available information on the function of indoleamine-2,3 dioxygenase enzyme in the tumor microenvironment.

Inhibitory Effects of Euphorbia tirucalli Lineu (Euphorbiaceae) Diluted Latex on Human and Canine Melanoma Cells.

Euphorbia tirucalli Lineu (Euphorbiaceae) is a tropical and subtropical ornamental and toxic plant. E. tirucalli produces a latex that is commonly used to treat neoplasms. This study aimed to evaluate the effects of diluted E. tirucalli latex (DETL) on human (SK-MEL-28) and canine (CBMY) melanoma cells. SK-MEL-28 (3 × 103 cells/well) and CBMY (6 × 103 cells/well) were cultivated in 96-well plates. The cells were treated with 50 µl/well of dilutions (1/2, 1/4, 1/8, 1/16, 1/32, 1/64, 1/128, 1/256, and 1/512) of a standard solution containing 1 mg/mL of the E. tirucalli latex (ETL) in DMEM. Control group cells received 50 µl/well of DMEM. After 24, 48, and 72 h of treatment, cell viability was assessed by the MTT assay. There was a significant decrease in viability at 48 and 72 hours after treatment for human melanoma cells and at 24, 48, and 72 hours for canine cells, mainly in higher dilutions of ETL. Human melanoma cells presented a typical U shape curve, characteristic of hormesis. To our knowledge, this is the first study showing inhibitory effects of DETL on canine melanoma cells. Therefore, DETL is a potentially new antineoplastic drug.

Development of a Sensitive Real-Time Fast-qPCR Based on SYBR® Green for Detection and Quantification of Chicken Parvovirus (ChPV).

Many viruses have been associated with runting and stunting syndrome (RSS). These viral infections mainly affect young chickens, causing apathy, depression, ruffled feathers, cloacal pasting, and diarrhea. Chicken Parvovirus (ChPV) is such an infection and has been detected in chickens showing signs of enteric diseases worldwide. Therefore, the present study aims to develop a sensitive real-time fast-qPCR assay based on SYBR® Green for detection and quantification of ChPV. A 561-bp non-structural (NS) gene was amplified and cloned, and a pair of primers was designed based on conserved nucleotide sequences on the NS gene of ChPV, the intercalating DNA reagent SYBR® Green was employed, and the Fast mode of a thermocycler was used. The assay detects 108 to 10¹ copies of the genome (CG). The limit of detection (LoD) was estimated to five CG, and the limit of quantification (LoQ) was estimated at ten CG. The standard curve efficiency was 101.94%, and the melting curve showed a unique clean peak and a melting temperature of 79.3 °C. The assay was specific to amplify the ChPV NS gene, and no amplification was shown from other viral genomes or in the negative controls. A total of 141 samples were tested using the assay, of which 139 samples were found positive. The highest CG value of ChPV was 5.7 × 106 CG/uL of DNA without apparent clinical signs of enteric disturbance, and 4.6 × 106 CG/uL DNA were detected in chickens with RSS.

Connexins and pannexins in liver damage.

Connexins and pannexins are key players in the control of cellular communication and thus in the maintenance of tissue homeostasis. Inherent to this function these proteins are frequently involved in pathological processes. The present paper reviews the role of connexins and pannexins in liver toxicity and disease. As they act both as sensors and effectors in these deleterious events connexins and pannexins could represent a set of novel clinical diagnostic biomarkers and drug targets.

Involvement of connexin43 in acetaminophen-induced liver injury.

BACKGROUND AND AIMS: Being goalkeepers of liver homeostasis, gap junctions are also involved in hepatotoxicity. However, their role in this process is ambiguous, as gap junctions can act as both targets and effectors of liver toxicity. This particularly holds true for drug-induced liver insults. In the present study, the involvement of connexin26, connexin32 and connexin43, the building blocks of liver gap junctions, was investigated in acetaminophen-induced hepatotoxicity. METHODS: C57BL/6 mice were overdosed with 300mg/kg body weight acetaminophen followed by analysis of the expression and localization of connexins as well as monitoring of hepatic gap junction functionality. Furthermore, acetaminophen-induced liver injury was compared between mice genetically deficient in connexin43 and wild type littermates. Evaluation of the toxicological response was based on a set of clinically relevant parameters, including protein adduct formation, measurement of alanine aminotransferase activity, cytokines and glutathione. RESULTS: It was found that gap junction communication deteriorates upon acetaminophen intoxication in wild type mice, which is associated with a switch in mRNA and protein production from connexin32 and connexin26 to connexin43. The upregulation of connexin43 expression is due, at least in part, to de novo production by hepatocytes. Connexin43-deficient animals tended to show increased liver cell death, inflammation and oxidative stress in comparison with wild type counterparts. CONCLUSION: These results suggest that hepatic connexin43-based signaling may protect against acetaminophen-induced liver toxicity.

Connexin32: a mediator of acetaminophen-induced liver injury?

Connexin32 is the building block of hepatocellular gap junctions, which control direct intercellular communication and thereby act as goalkeepers of liver homeostasis. This study was set up to investigate whether connexin32 is involved in hepatotoxicity induced by the analgesic and antipyretic drug acetaminophen. To this end, whole body connexin32 knock-out mice were overdosed with acetaminophen followed by sampling at different time points within a 24-h time frame. Evaluation was done based upon a series of clinically and mechanistically relevant read-outs, including protein adduct formation, histopathological examination, measurement of alanine aminotransferase activity, cytokine production, levels of reduced and oxidized glutathione and hepatic protein amounts of proliferating cell nuclear antigen. In essence, it was found that genetic ablation of connexin32 has no influence on several key events in acetaminophen-induced hepatotoxicity, including cell death, inflammation or oxidative stress, yet it does affect production of protein adducts as well as proliferating cell nuclear antigen steady-state protein levels. This outcome is not in line with previous studies, which are contradicting on their own, as both amplification and alleviation of this toxicological process by connexin32 have been described. This could question the suitability of the currently available models and tools to investigate the role of connexin32 in acetaminophen-triggered hepatotoxicity.

Comparative Aspects of Canine Melanoma.

Melanomas are malignant neoplasms originating from melanocytes. They occur in most animal species, but the dog is considered the best animal model for the disease. Melanomas in dogs are most frequently found in the buccal cavity, but the skin, eyes, and digits are other common locations for these neoplasms. The aim of this review is to report etiological, epidemiological, pathological, and molecular aspects of melanomas in dogs. Furthermore, the particular biological behaviors of these tumors in the different body locations are shown. Insights into the therapeutic approaches are described. Surgery, chemotherapy, radiotherapy, immunotherapy, and the outcomes after these treatments are presented. New therapeutic perspectives are also depicted. All efforts are geared toward better characterization and control of malignant melanomas in dogs, for the benefit of these companion animals, and also in an attempt to benefit the treatment of human melanomas.

The role of prolactin in prostatic inflammation / O papel da prolactina na inflamação prostática

Several reports have shown that prolactin (PRL) plays a role in prostatic growth, but few studies considered the role of PRL in the process of prostatic inflammation. Young (45 ± 5 days old) and adult (75 ± 5 days old) male Wistar rats were subcutaneously injected daily with domperidone (4.0 mg.kg-1) to maintain high serum PRL levels. The animals were treated for 15, 30, 45 or 60 days. Blood and prostate samples were collected at the end of each treatment for PRL dosage and histological analysis, respectively. Only young animals treated with DOMP for 15 and 30 days displayed inflammatory infiltrate in the prostate. These results confirm literature data in regards to PRL involvement in inducing prostate inflammation. Moreover, it was concluded that young animals are more susceptible then adults to the PRL action concerning prostate inflammation...

A prolactina (PRL) influencia o crescimento prostático, entretanto poucos estudos investigaram o papel da PRL na inflamação prostática. Ratos Wistar jovens (45 ± 5 dias de idade) e adultos (75 ± 5 dias de idade) receberam injeções subcutâneas diárias de domperidona (4,0 mg.kg-1) para manter níveis séricos altos de PRL. Os animais foram tratados por 15, 30, 45 ou 60 dias. Amostras de sangue e próstata foram coletadas ao final dos tratamentos para dosagem de PRL e análise histológica, respectivamente. Apenas os animais jovens tratados com domperidona por 15 e 30 dias apresentaram infiltrado inflamatório na próstata. Esses resultados confirmaram a participação da PRL na indução da inflamação prostática. A conclusão obtida foi que animais jovens são mais susceptíveis à ação da PRL na inflamação da próstata que os adultos...

Paullinia cupana Mart. var. sorbilis, guarana, increases survival of Ehrlich ascites carcinoma (EAC) bearing mice by decreasing cyclin-D1 expression and inducing a G0/G1 cell cycle arrest in EAC cells.

The objective of this work is to report the antiproliferative effect of P. cupana treatment in Ehrlich Ascites Carcinoma (EAC)-bearing animals. Female mice were treated with three doses of powdered P. cupana (100, 1000 and 2000 mg/kg) for 7 days, injected with 10(5) EAC cells and treated up to day 21. In addition, a survival experiment was carried out with the same protocol. P. cupana decreased the ascites volume (p = 0.0120), cell number (p = 0.0004) and hemorrhage (p = 0.0054). This occurred through a G1-phase arrest (p < 0.01) induced by a decreased gene expression of Cyclin D1 in EAC cells. Furthermore, P. cupana significantly increased the survival of EAC-bearing animals (p = 0.0012). In conclusion, the P. cupana growth control effect in this model was correlated with a decreased expression of cyclin D1 and a G1 phase arrest. These results reinforce the cancer therapeutic potential of this Brazilian plant.

Clinical study of cryosurgery efficacy in the treatment of skin and subcutaneous tumors in dogs and cats.

OBJECTIVE: To evaluate the efficacy of cryosurgery for treatment of skin and subcutaneous tumors in dogs and cats. STUDY DESIGN: Prospective study. ANIMALS: Dogs (n=20), cats (10). METHODS: Cutaneous or subcutaneous tumors were treated by liquid nitrogen cryosurgical spray (1 cm from target tissue at 90 degrees until a 5-mm halo of frozen tissue was achieved) for 15-60 seconds. Malignant lesions had 3 freeze-thaw cycles benign tumors, 2 cycles. The second or third freeze cycle was performed after complete thaw of the preceding freeze. Wounds healed by second intention. Follow-up was weekly for 1 month and then twice monthly until wounds healed, and final outcome was determined by telephone interview of owners. RESULTS: Tumor size ranged from 0.3 to 11 cm diameter with 28 (60%) being 0.3-1 cm; 8 (17%) 1.1-3 cm, and 11 (23%) >3.4 cm. Complications included edema, erythema and for extremity lesions, pain and lameness. Treated lesions (n=47) had an overall remission of 98% (mean follow-up, 345+/-172.02 days [range, 150-750 days]). One malignant peripheral nerve sheath tumor recurred 7 months after cryosurgical treatment. CONCLUSION: Cryosurgery is an efficient method for treatment of skin and subcutaneous tumors in dogs and cats. CLINICAL RELEVANCE: Cryosurgical ablation is an effective means of treating small cutaneous or subcutaneous tumors in dogs and cats, especially in older animals where wound closure or cosmetic outcome might limit surgical excision alone.

Defective gap junctional intercellular communication in the carcinogenic process.

Gap junctions are membrane structures made of intercellular channels which permit the diffusion from cytoplasm to cytoplasm of small hydrophilic molecules. Nearly 40 years ago, the loss of functional gap junctions has been described in cancer cells and led to the hypothesis that such type of intercellular communication is involved in the carcinogenesis process. From this time, a lot of data has been accumulated confirming that gap junctions are frequently decreased or absent in cancer cells whatever their tissue and species origins. Here, we review such data by insisting on the possible links existing between altered gap-junctional intercellular communication capacity (or the altered expression of their constitutive proteins, the connexins) and the stages of cancer progression in various cancer models. Then, we analyse particular aspects of the disturbance of connexin-mediated communication in cancer such as the cytoplasmic localization of connexins, the lack of heterologous communication between cancer cells and normal cells, the role of connexin gene mutations in cancer. In a separate part of the review, we also analyse the disturbance of gap-junctional intercellular communication during the late stages of cancer (invasion and metastasis processes).

Clinical evaluation of random skin flaps based on the subdermal plexus secured with sutures or sutures and cyanoacrylate adhesive for reconstructive surgery in dogs.

OBJECTIVE: To evaluate the use of subdermal plexus skin flaps for closing defects after excision of cutaneous and subcutaneous tumors in dogs and to compare outcome of flaps secured with sutures and those secured with butyl-cyanoacrylate and intermittent sutures. STUDY DESIGN: Clinical study. ANIMALS: Fifteen dogs. METHODS: After excision of cutaneous or subcutaneous tumors the skin defect was reconstructed by random flaps based on the subdermal plexus. Flap skin edges were apposed with simple interrupted 4-0 monofilament nylon sutures (group 1; 5 dogs) or nylon sutures alternated with butyl-cyanoacrylate adhesive (group 2; 10 dogs). Flaps were evaluated every 48 hours when bandages were changed, until complete healing. RESULTS: Random flaps based on the subdermal plexus were effectively used to close wound defects; mean flap survival was 89%. Partial flap necrosis occurred in 4 dogs. Wound margins apposed with butyl-cyanoacrylate had thinner and more esthetic scars than sutured margins. CONCLUSION: Random flaps based on the subdermal plexus proved to be versatile for covering limb wounds after excision of cutaneous or subcutaneous tumors. Mean survival rate was comparable to that reported for axial pattern flaps. Butyl-cyanoacrylate adhesive was easy to apply, allowed accurate margin apposition with good cosmetic outcome and reduced sutures needed. CLINICAL RELEVANCE: Cyanoacrylate adhesive should be considered in lieu of suture closure to secure random skin flaps based on the subdermal plexus in dogs.

Effect of Pfaffia paniculata (Brazilian ginseng) on the Ehrlich tumor in its ascitic form.

The roots of Pfaffia paniculata (Brazilian ginseng) have been indicated for the treatment of several diseases, among which the cancer. The purpose of this study was to investigate experimentally the possible antineoplastic effect of this root. Firstly, a toxicity study was performed in which the doses of 400 and 200 mg/Kg of the powdered root were administered by gavage for 10 days to BALB/cICB mice. The mice did not lose weight during the treatment. No increase in serum alanine-aminotransferase neither histopathological alteration (liver, kidney and spleen) was observed in mice treated with P. paniculata. The effect of this root on the ascitic Ehrlich tumor in BALB/cICB mice was then investigated. Male mice received, by gavage, once a day, 200 mg/Kg of the powdered root of P. paniculata or distilled water, as control, for 20 days. This protocol started 10 days before tumor inoculation with 5 x 10(6) cells i.p., and lasted until 10 days after. The ascitic tumor was evaluated by the quantification of the volume of the ascitic fluid, relative number of tumor cells and total number of tumor cells. A decrease in the total ascitic volume was observed in P. paniculata treated mice, that was followed by a numerical decrease in the total number of Ehrlich tumor cells. These results may indicate that P. paniculata anti-inflammatory effects were responsible by the decrease in the total ascitic fluid. In addition, the presence of tumor-cell inhibitory factors in P. paniculata roots is in agreement with other in vitro studies. The mechanisms of such tumor inhibition should be further investigated.