Treatment of chronic immunomediated thrombocytopenia with mesenquimal stem cells in dog: case report / Tratamento de trombocitopenia imunomediada crônica com células-tronco mesenquimais em cão: relato de caso

Immunomediated thrombocytopenia is a systemic metabolic disorder in which the platelet count falls below reference values, as the patient's immune system destroys them. The main clinical signs in thrombocytopenia are petechial, hemorrhages, ecchymoses and suffusions. Hematomas can also occur in coagulation disorders. The diagnosis is based on clinical findings and hematological examinations. The treatment consists of the use of corticosteroids and immunosuppressants, delaying cell destruction, and may last for months, not always obtaining a cure for the disease. The present work reports the use of therapy with allogeneic mesenchymal stem cells, derived from the adipose tissue of dogs, for the treatment of chronic immunomediated thrombocytopenia, with an evolution of more than one year, in a Pinscher dog. The alternative treatment showed a good evolution, keeping platelets within the reference values during the treatment, giving the patient quality of life and removing the need for continuous medication for homeostasis after treatment.

A trombocitopenia imunomediada é uma desordem metabólica sistêmica, na qual a contagem plaquetária fica abaixo dos valores de referência, pois o sistema imunológico do paciente a destrói. O principal sinal clínico na trombocitopenia são hemorragias, petequiais, equimoses e sufusões. Hematomas podem ocorrer também em alterações da coagulação. O diagnóstico baseia-se nos achados clínicos e nos exames hematológicos. O tratamento consiste na utilização de corticosteroides e imunossupressores, o que retarda a destruição celular, mas pode se prolongar por meses, nem sempre obtendo cura da doença. O presente trabalho relata a utilização da terapia com células-tronco mesenquimais alogênicas, oriundas do tecido adiposo de cães, para tratamento de trombocitopenia imunomediada crônica, com evolução de mais de um ano, em um cão da raça Pinscher. O tratamento alternativo revelou boa evolução, pois manteve as plaquetas dentro dos valores de referência durante o tratamento, o que proporcionou qualidade de vida ao paciente e tornou desnecessárias medicações de uso contínuo para a homeostase após o tratamento.

Modulation of periovulatory endocrine profiles in beef cows: consequences for endometrial glucose transporters and uterine fluid glucose levels.

In beef cattle, proestrus estradiol and subsequent progesterone (P4) concentrations can regulate the endometrial characteristics and thereby determine maternal receptivity toward the embryo. However, the underlying mechanisms linking periovulatory endocrine profiles to receptivity, which is crucial to obtain pregnancy, need to be elucidated. We hypothesized that the size of the preovulatory follicle (POF) and subsequent circulating P4 concentrations, during early diestrus, modulate endometrial levels of glucose transporter transcripts and proteins, and subsequently affect the luminal glucose availability in the uterus. Therefore, follicle growth of Nelore cows was manipulated, and cows were assigned to 2 experimental groups: (1) large follicle and large corpus luteum (LF-LCL) group with a large POF and corpus luteum (CL); and (2) small follicle and small corpus luteum (SF-SCL) group with a small POF and CL. At day 7 post gonadotropin-releasing hormone induced ovulation (gonadotropin-releasing hormone treatment = day 0), animals were slaughtered (n = 18 per group), and uterine tissues and washings were collected for characterization of glucose transporters and glucose levels, respectively. The diameter of POF was larger (P < 0.05) in the LF-LCL cows compared with their SF-SCL counterparts (12.8 ± 0.4 vs 11.1 ± 0.4 mm). Furthermore, CL size (17.49 ± 0.88 vs 14.48 ± 0.52 mm) and circulating P4 concentrations at day 7 (4.5 ± 1.0 vs 3.3 ± 1.1 ng/mL, P < 0.05) were significantly higher in the LF-LCL cows compared with the SF-SCL cows. No differences (P > 0.05) were detected in gene expression patterns of SLC2A1, SLC2A3, SLC2A4, SLC2A5, SLC5A1, ATP1A2, ATP1B2, and SLC37A4. However, the protein abundance of endometrial SLC2A1was increased in the LF-LCL group compared with the SF-SCL group (P < 0.05). SLC2A1 and SLC2A4 protein products were mainly identified at the endometrial luminal and glandular epithelium membranes as well as in the endometrial stroma. Glucose concentrations in uterine washings were similar between groups. In conclusion, we provided information on the potential link between endocrine profiles and glucose transport pathways in the bovine endometrium. More specifically, our data reveal that the size of the POF, and subsequent P4 concentrations, do not functionally affect the main endometrial glucose transporter pathways or uterine fluid glucose concentrations during diestrus.

Storage of bovine reproductive tissues and RNA extracts on ice for 24 h or repeated freeze-thaw cycles do not affect RNA integrity.

The aims of this study were to test (i) the effect of time of tissue and RNA extracts storage on ice and (ii) the effect of repeated freeze-thaw cycles on RNA integrity and gene expression of bovine reproductive tissues. Fragments of endometrium (ENDO), corpus luteum (CL) and ampulla (AMP) were subdivided and incubated for 0, 1, 3, 6, 12 or 24 h on ice. RNA extracts were incubated on ice for 0, 3, 12 or 24 h, or exposed to 1, 2, 4 or 6 freeze-thaw cycles. RNA integrity number (RIN) was estimated. Expression of progesterone receptor (PGR) and cyclophilin genes from RNA extracts stored on ice for 0 or 24 h, and 1 or 6 freeze-thaw cycles was measured by qPCR. Tissue and RNA extract incubation on ice, and repeated freeze-thaw cycles did not affect RIN values of RNA from ENDO, CL or AMP. Storage on ice or exposure to freeze-thaw cycles did not affect Cq values for PGR or cyclophilin genes. In conclusion, neither generalized RNA degradation nor specific RNA degradation was affected by storage of tissue or RNA extracts on ice for up to 24 h, or by up to 6 freeze-thaw cycles of RNA extracts obtained from bovine ENDO, CL and AMP.

The role of proestrus on fertility and postovulatory uterine function in the cow

Modern protocols to synchronize ovulations for timed artificial insemination and timed embryo transfer that include manipulations in the proestrus period (i.e., between luteolysis and estrus) affect fertility in cattle. Specifically, stimulating pre-ovulatory follicle growth and exposure to estrogens after CL regression increase the proportion of cows pregnant and decrease late embryo mortality. Such effects may be due to both pre-ovulatory actions of estrogens and post-ovulatory actions of progesterone, as concentrations of the later hormone may be changed in response to manipulations conducted during proestrus. In the first portion of this paper we describe strategies used recently to manipulate the proestrus period in protocols for synchronization of ovulation, and to present evidence of their effects on fertility. Manipulations of timing and prominence of sex steroids during the proestrus and early diestrus that affect fertility may act on targets such as the endometrium. This tissue expresses receptors for both estrogens and progesterone and these hormones change endometrial function to support conceptus growth and pregnancy maintenance. However, specific cellular and molecular mechanisms through which fertility is affected via manipulations of the proestrus are poorly understood. In the second portion of this paper we describe a well-defined animal model to study changes in endometrial function induced by manipulations conducted during the proestrus. Such manipulations induced endometrial changes on sex steroid receptors expression, cell proliferation, oxidative metabolism and eicosanoid synthesis in the uterus, but not on glucose transport to uterine lumen . In summary, evidence is accumulating to support a positive role of increasing duration and estrogen availability during the proestrus on fertility to synchronization protocols. Such positive effects may be through changes in endometrial function to stimulate conceptus growth and survival. (AU)

The role of proestrus on fertility and postovulatory uterine function in the cow

Modern protocols to synchronize ovulations for timed artificial insemination and timed embryo transfer that include manipulations in the proestrus period (i.e., between luteolysis and estrus) affect fertility in cattle. Specifically, stimulating pre-ovulatory follicle growth and exposure to estrogens after CL regression increase the proportion of cows pregnant and decrease late embryo mortality. Such effects may be due to both pre-ovulatory actions of estrogens and post-ovulatory actions of progesterone, as concentrations of the later hormone may be changed in response to manipulations conducted during proestrus. In the first portion of this paper we describe strategies used recently to manipulate the proestrus period in protocols for synchronization of ovulation, and to present evidence of their effects on fertility. Manipulations of timing and prominence of sex steroids during the proestrus and early diestrus that affect fertility may act on targets such as the endometrium. This tissue expresses receptors for both estrogens and progesterone and these hormones change endometrial function to support conceptus growth and pregnancy maintenance. However, specific cellular and molecular mechanisms through which fertility is affected via manipulations of the proestrus are poorly understood. In the second portion of this paper we describe a well-defined animal model to study changes in endometrial function induced by manipulations conducted during the proestrus. Such manipulations induced endometrial changes on sex steroid receptors expression, cell proliferation, oxidative metabolism and eicosanoid synthesis in the uterus, but not on glucose transport to uterine lumen . In summary, evidence is accumulating to support a positive role of increasing duration and estrogen availability during the proestrus on fertility to synchronization protocols. Such positive effects may be through changes in endometrial function to stimulate conceptus growth and survival.