Establishment of effective and safe recipient preparation for germ-cell transplantation with intra-testicular busulfan treatment in pre-pubertal Barbari goats.

The busulfan, an alkylating agent, suppresses endogenous spermatogenesis in recipient testes. However, considering a wide variation in the effects of busulfan among animal species, its dosage and route of infusion need optimization to prepare effective and safe recipients. Thus, the current study aimed to create a suitable recipient goat model for germ cell (Gc) transplantation through a single intra-testicular (i.t.) busulfan infusion under ultrasonographic (USG) guidance. As observed through the infusion of trypan blue under USG guidance into mediastinum testis (MT) of pre-pubertal Barbari bucks, 3-5 mL of trypan blue solution could fill almost 80% of seminiferous tubules. Thereafter, in Experiment-1, the effect of different busulfan doses (mg/kg) i.e. 0 [negative control, Group (Gr) 1; 0 mg/kg-MT], 1 (Gr 2; 1 mg/kg-MT), 2 (Gr 3; 2 mg/kg-MT), and 3 (Gr 4; 3 mg/kg-MT) were studied. Further, in Experiment-2, sterilizing effects of busulfan infusion through two different routes [MT or cavum vaginale (CV)] were compared. Following i.t. busulfan treatment, no adverse physiological effects or body weight loss were detected. The histological analyses demonstrate a dose-dependent depletion of Gc with almost complete loss of Gc and spermatogenic activities in Gr 3 and 4, and extensive fibrosis in Gr 4. A considerable suppression of spermatogenesis marked with devoid of endogenous spermatogonial population and absence of significant (P > 0.05) effect on key hematological variables were observed in 2 mg/kg-MT Gr. These findings coupled with the results of significant (P < 0.05) down-regulation of marker genes of undifferentiated spermatogonia (THY-1 and PLZF), Gc pluripotency (UCHL-1, OCT-4, and DDX-4), and adhesion (E-cadherin and ß-integrin); up-regulation of apoptotic genes (ID - 4 and BCL-6), and unchanged expression of Sertoli cell marker (vimentin), confirmed the safe and efficient depletion of endogenous Gc in 2 mg/kg-MT Gr. Furthermore, the effect of busulfan infusion on scrotal-testicular biometry, endocrine variables (plasma cortisol and testosterone), and Gc removal was more evident when busulfan was infused into MT than into CV. Overall, the results demonstrated that 2.0 mg/kg is an optimal single dose of busulfan when infused into the MT under USG guidance for the preparation of pre-pubertal recipient bucks. Overall, this study provides a basis to prepare suitable recipients through providing an available niche for efficient Gc transplantation in goats.

Germ Cell Transplantation in Stallion Testes.

The production of donor-derived sperm using spermatogonial stem cell transplantation has been studied in various animals including mice, rats, goats, boar, dogs, sheep, and monkeys. However, germ cell transplantation has not been applied in stallions. The objective of this study was to produce donor germ cell-derived sperm using germ cell transplantation in stallions. Donor germ cells were transplanted into the parenchyma of 3 recipient stallions that had been treated with busulfan IV injections of 15 mg/kg body weight. For the preparation of donor single germ cells, tissue (20 g) from each testis was subjected to a 2-enzyme digestion procedure. Donor testicular germ cells in minimum essential medium α supplemented with 10% fetal bovine serum were transplanted in the testis of recipient stallions at a rate of 2 ml/min. The semen of each recipient stallion was collected using an artificial vagina at 8 weeks after germ cell transplantation. General sperm evaluation and libido tests were performed. Microsatellite fingerprinting with 17 markers was performed to identify the presence of donor-derived sperm in the semen of the recipient stallions. Sperm were observed to have total and progressive motility exceeding 50% throughout the experimental period. The libido of the recipient stallions was unchanged. No donor-derived sperm could be detected in the semen of the recipient stallions by genotyping. In conclusion, the transplantation of donor germ cells into the testicular parenchyma of stallions was not an optimal transplantation technique for producing donor-derived sperm.

Assessment of Yangtze sturgeon as recipient for the production of American paddlefish gametes through spermatogonia transplantation.

The Chinese paddlefish (Psephurus gladius), one of the world's largest freshwater fish, was last seen alive in 2003; they are presumed now to be extinct. In fish, germ cell transplantation is currently known as one of the most powerful assisted reproductive technologies for the conservation of endangered species. In the event that a Chinese paddlefish is unexpectedly caught in the near future, we aimed to develop an experimental strategy to produce paddlefish gametes in the gonads of surrogate sturgeon. Spermatogonia were collected from the testes of 2.5-year-old immature male American paddlefish (Polyodon spathula), the species most closely related to the Chinese paddlefish, by Percoll gradient centrifugation, and transplanted into the peritoneal cavity of Yangtze sturgeon (Acipenser dabryanus) larvae at 7-8 days post-hatch. At two months post-transplantation, donor-derived spermatogonia had efficiently colonized in the recipient gonads and proliferated. A PCR analysis developed to detect xenogenic donor-derived mtDNA sequences in recipient gonads revealed that American paddlefish germ cells survived for at least seven months after transplantation in the gonads of Yangtze sturgeon recipients. These results show that the somatic microenvironment of Yangtze sturgeon gonads was able to support the colonization, proliferation, and survival of xenogeneic germ cells from a different taxonomic family. This study provides key information that could lead to future restoration of Chinese paddlefish using germ cell transplantation.

Efficacy of autologous bone marrow mononuclear cell transplantation in dogs with chronic spinal cord injury.

Background: Spinal cord injury (SCI) is relatively common in dogs and is a devastating condition involving loss of sensory neurons and motor neurons. However, the main clinical protocol for the management of SCI is surgery to decompress and stabilize the vertebra. Cell transplantation therapy is a very promising strategy for the treatment of chronic SCI, but extensive preclinical and clinical research work remains. Aim: The aim of this study is to confirm the effect of bone marrow-derived mononuclear cell (BM-MNC) transplantation for chronic SCI in dogs. Methods: We tested the treatment efficiency of chronic SCI in 12 dogs using BM-MNC transplantation. Neurological evaluation used the Texas Spinal Cord Injury Scale (TSCIS). Concurrently, we characterized the transplanted cells by evaluation using quantitative real-time polymerase chain reaction, flow cytometry, and enzyme-linked immunosorbent assay. Result: All dogs had a pre-transplantation TSCIS score of 0. Two animals did not show any improvement in their final TSCIS scores. The remaining 10 dogs (83.4%) achieved improvement in the final TSCIS scores. Five of them (41.7%) regained ambulatory function with a TSCIS score greater than 10. We determined that canine BM-MNCs expressed hepatocyte growth factor (HGF) mRNA at higher levels than other cytokines, with significant increases in HGF levels in cerebrospinal fluid within 48 hours after autologous BM-MNC transplantation into the subarachnoid space of the spinal dura matter in dogs. Conclusions: BM-MNC transplantation may be effective for at least some cases of chronic SCI.

Aging- and temperature-related activity of spermatogonial stem cells for germ cell transplantation in medaka.

Spermatogonial transplantation can contribute to developing a novel method of producing seedlings for both aquaculture and biotic conservation. This study's purpose was to investigate aging- and temperature-related changes in the numbers and stem cell functions of type-A spermatogonia (ASG) in the model fish medaka (Oryzias latipes). The ASG numbers in medaka of different ages were quantified via histological observation and enzymatic dissociation of vasa-Gfp medaka testes. The ASG numbers were higher in eight-month-old medaka (maturation) than in four-month-old medaka (the onset of maturation). However, ASG numbers decreased in 18-month-old medaka (senescence). Low water temperature appeared to slow down both testis development and aging processes. To study the effects of aging on ASG stem cell activity, testicular cell suspensions containing GFP-expressed ASG were prepared from vasa-Gfp medaka donors at 4 and 18 months of age and transplanted into recipient hybrid larvae of medaka (O. latipes x O. curvinotus), which provided young stem-cell-niches. The findings revealed no significant differences in ASG colonization rates isolated from medaka of different ages. Each group displayed similar rates of germ-line transmission. Furthermore, water temperature had no significant effects on each ASG's stem cell activity. Taken together, these results indicated that aging and temperature affect ASG numbers. However, ASG isolated from medaka with different ages were transplanted into gonads with a young niche microenvironment, and there was no evidence of donor aging on stem cell activity.

Cell engraftment, vascularization, and inflammation after treatment of equine distal limb wounds with endothelial colony forming cells encapsulated within hydrogel microspheres.

BACKGROUND: Endothelial colony forming cells (ECFCs) may be useful therapeutically in conditions with poor blood supply, such as distal limb wounds in the horse. Encapsulation of ECFCs into injectable hydrogel microspheres may ensure cell survival and cell localization to improve neovascularization and healing. Autologous ECFCs were isolated from 6 horses, labeled with quantum nanodots (QD), and a subset were encapsulated in poly(ethylene) glycol fibrinogen microspheres (PEG-Fb MS). Full-thickness dermal wounds were created on each distal limb and injected with empty PEG-Fb MS, serum, ECFCs, or ECFCs encapsulated into PEG- Fb MS (ECFC/MS). Analysis included wound surface area (WSA), granulation tissue scoring (GS), thermography, collagen density staining, and immunohistochemical staining for endothelial and inflammatory cells. The purpose of this study was to track cell location and evaluate wound vascularization and inflammatory response after injection of ECFC/MS or naked ECFCs in equine distal limb wounds. RESULTS: ECFCs were found near and within newly formed blood vessels up to 3 weeks after injection. ECFC and ECFC/MS groups had the greatest blood vessel quantity at week 1 in the wound periphery. Wounds treated with ECFCs and ECFC/MS had the lowest density of neutrophils and macrophages at week 4. There were no significant effects of ECFC or ECFC/MS treatment on other measured parameters. CONCLUSIONS: Injection of microsphere encapsulated ECFCs was practical for clinical use and well-tolerated. The positive ECFC treatment effects on blood vessel density and wound inflammation warrant further investigation.

Donor sperm production in heterologous recipients by testis germ cell transplantation in the dromedary camel.

The object of this study was to investigate if testis germ cell transplantation (TGCT) into a heterologous recipient would result in donor-origin spermatogenesis in the dromedary camel. First, we investigated a workable protocol for TGCT in camels, including donor cell isolation, enrichment by density gradient centrifugation (Percoll and Bovicoll), rete testis injection and microsatellite detection of donor and recipient genotypes. Second, the effects of three doses of Dolichos biflorus agglutinin (DBA), a glycoprotein that specifically binds to gonocytes or Type A spermatogonia, on testis germ cell depletion were investigated by direct injection into the rete testis of a male camel. Seven recipients were prepared with DBA treatment, two males were castrated at 4 weeks for depletion assessment and the remaining five received donor cells 4-6 weeks after treatment. On average, ~17 million cells were isolated per gram of testis tissue, with 19.5±1.9% DBA-positive (DBA+) cells. Percoll centrifugation yielded a 1.5-fold increase in DBA+ cells while Bovicoll centrifugation produced a 2.5-fold increase from the input cells of 18.6±2.1% DBA+ cells. Semen was collected from the recipients 13-20 weeks after transfer and the presence of donor DNA in the samples was determined using microsatellite markers. In two of the five recipients, all semen samples were shown to be positive for donor-derived cells. These results demonstrate for the first time that: (1) heterologous testicular germ cell transplantation in camels is feasible and the recipients are able to produce spermatozoa of donor origin and (2) DBA can be used effectively to deplete endogenous stem cells.

Interspecific germ cell transplantation: a new light in the conservation of valuable Balkan trout genetic resources?

Interspecific transplantation of germ cells from the brown trout Salmo trutta m. fario and the European grayling Thymallus thymallus into rainbow trout Oncorhynchus mykiss recipients was carried out in order to improve current practices in conservation of genetic resources of endangered salmonid species in the Balkan Peninsula. Current conservation methods mainly include in situ efforts such as the maintenance of purebred individuals in isolated streams and restocking with purebred fingerlings; however, additional ex situ strategies such as surrogate production are needed. Steps required for transplantation such as isolation of high number of viable germ cells and fluorescent labeling of germ cells which are to be transplanted have been optimized. Isolated and labeled brown trout and grayling germ cells were intraperitoneally transplanted into 3 to 5 days post hatch rainbow trout larvae. Survival of the injected larvae was comparable to the controls. Sixty days after transplantation, fluorescently labeled donor cells were detected within the recipient gonads indicating successful incorporation of germ cells (brown trout spermatogonia and oogonia-27%; grayling spermatogonia-28%; grayling oogonia-23%). PCR amplification of donor mtDNA CR fragments within the recipient gonads additionally corroborated the success of incorporation. Overall, the transplantation method demonstrated in this study presents the first step and a possible onset of the application of the germ cell transplantation technology in conservation and revitalization of genetic resources of endangered and endemic species or populations of salmonid fish and thus give rise to new or improved management strategies for such species.

Testicular germ line cell identification, isolation, and transplantation in two North American catfish species.

Our aim was to transplant blue catfish germ line stem cells into blastulae of triploid channel catfish embryos to produce interspecific xenogenic catfish. The morphological structure of the gonads of blue catfish (Ictalurus furcatus) in ~ 90- to 100-day-old juveniles, two-year-old juveniles, and mature adults was studied histologically. Both oogonia (12-15 µm, diameter with distinct nucleus 7-8 µm diameter) and spermatogonia (12-15 µm, with distinct nucleus 6-7.5 µm diameter) were found in all ages of fish. The percentage of germ line stem cells was higher in younger blue catfish of both sexes. After the testicular tissue was trypsinized, a discontinuous density gradient centrifugation was performed using 70, 45, and 35% Percoll to enrich the percentage of spermatogonial stem cells (SSCs). Four distinct cell bands were generated after the centrifugation. It was estimated that 50% of the total cells in the top band were type A spermatogonia (diameter 12-15 µm) and type B spermatogonia (diameter 10-11 µm). Germ cells were confirmed with expression of vasa. Blastula-stage embryos of channel catfish (I. punctatus) were injected with freshly dissociated blue catfish testicular germ cells as donor cells for transplantation. Seventeen days after the transplantation, 33.3% of the triploid channel catfish fry were determined to be xenogenic catfish. This transplantation technique was efficient, and these xenogenic channel catfish need to be grown to maturity to verify their reproductive capacity and to verify that for the first time SSCs injected into blastulae were able to migrate to the genital ridge and colonize. These results open the possibility of artificially producing xenogenic channel catfish males that can produce blue catfish sperm and mate with normal channel catfish females naturally. The progeny would be all C × B hybrid catfish, and the efficiency of hybrid catfish production could be improved tremendously in the catfish industry.

Establishment of intraperitoneal germ cell transplantation for critically endangered Chinese sturgeon Acipenser sinensis.

Recent progress in germ cell transplantation techniques in fish has paved the way for the conservation of endangered species. Here, we developed an intraperitoneal germ cell transplantation procedure using Chinese and Dabry's sturgeon as donor and recipient species, respectively. Histological analysis revealed that primordial germ cells migrated on the peritoneal wall at 16 days post-hatch (dph) in Dabry's sturgeon. The genital ridges of Dabry's sturgeon (recipient) first formed at 28 dph, suggesting that for successful colonization of donor germ cells in the recipient gonads, the transplantation should be performed earlier than this age. Sexual dimorphism of gonadal structure was first observed at 78 dph. Gonadal germ cell proliferation was not seen in either sex during this period. Immunohistochemistry using the anti-Vasa antibody found that donor testes from 2-year-old Dabry's sturgeon mainly consisted of single- or paired-type A spermatogonia, while donor ovaries from 11.5-year-old Chinese sturgeon had perinucleolus stage oocytes and clusters of oogonia. Donor cells isolated from Dabry's sturgeon testes or Chinese sturgeon ovary labeled with PKH26 fluorescent dye were transplanted into the peritoneal cavity of the 7- or 8-dph Dabry's sturgeon larvae. More than 90% and 70% of transplanted larvae survived after 2 days post-transplantation (dpt) and 51 dpt, respectively. At 51 dpt, PKH26-labeled cells exhibiting germ cell-specific nuclear morphology and diameter were observed in excised recipient gonads by fluorescent and confocal microscopy. The colonization rate of allogeneic testicular germ cell transplantation (Group 1) was 70%, while that of two batches of xenogeneic ovarian germ cell transplantation (Group 2 and Group 3) were 6.7% and 40%, respectively. The ratio of colonized germ cells to endogenous germ cells was 11.96%, 5.35% and 3.56% for Group 1, Group 2 and Group 3, respectively. Thus, we established a germ cell transplantation technique for the critically endangered Chinese sturgeon using the most closely related species as a recipient and demonstrated the successful preparation of transplantable female germ cells from aged adult Chinese sturgeon.

Establishment of oct4:egfp transgenic and oct4:egfp /ß-actin:DsRed double transgenic medaka lines.

As a model to examine cellular multipotency in fish, we established a medaka transgenic (Tg) Tru.oct4:egfp line carrying the green fluorescence protein (GFP) cDNA under control of the Takifugu rubripes oct4 promoter. In this Tg line, GFP could be used to examine both maternal and zygotic oct4 expression during embryogenesis. In addition, while adult Tg fish did not express GFP in any somatic cells, activation of GFP expression was initiated in regenerating fins after amputation. In vitro, some of the cell populations that migrated from fin explants expressed GFP, implying that GFP could be used to monitor oct4 expression in both embryos and in regenerating tissues in the Tru.oct4:egfp Tg line. Next, crossing with ß-actin:DsRed Tg line in which all cells emit red fluorescence by expression of red fluorescent protein (RFP) under the ß-actin promoter, we prepared a Tru.oct4:egfp /ß-actin:DsRed double Tg line. In the double Tg line, early embryonic cells were +GFP/+RFP double positive. In vitro fin cell culture, a small number of +GFP/+RFP double positive cells could be discriminated from other -GFP/+RFP cells. Thus, when transplanted into wild-type medaka, this double Tg line can be used to trace the fate of the transplanted cells using RFP fluorescence after the loss of GFP expression.

M1 and M2 macrophage recruitment during tendon regeneration induced by amniotic epithelial cell allotransplantation in ovine.

Recently, we have demonstrated that ovine amniotic epithelial cells (oAECs) allotransplanted into experimentally induced tendon lesions are able to stimulate tissue regeneration also by reducing leukocyte infiltration. Amongst leukocytes, macrophages (Mφ) M1 and M2 phenotype cells are known to mediate inflammatory and repairing processes, respectively. In this research it was investigated if, during tendon regeneration induced by AECs allotransplantation, M1Mφ and M2Mφ phenotype cells are recruited and differently distributed within the lesion site. Ovine AECs treated and untreated (Ctr) tendons were explanted at 7, 14, and 28 days and tissue microarchitecture was analyzed together with the distribution and quantification of leukocytes (CD45 positive), Mφ (CD68 pan positive), and M1Mφ (CD86, and IL12b) and M2Mφ (CD206, YM1 and IL10) phenotype related markers. In oAEC transplanted tendons CD45 and CD68 positive cells were always reduced in the lesion site. At day 14, oAEC treated tendons began to recover their microarchitecture, contextually a reduction of M1Mφ markers, mainly distributed close to oAECs, and an increase of M2Mφ markers was evidenced. CD206 positive cells were distributed near the regenerating areas. At day 28 oAECs treated tendons acquired a healthy-like structure with a reduction of M2Mφ. Differently, Ctr tendons maintained a disorganized morphology throughout the experimental time and constantly showed high values of M1Mφ markers. These findings indicate that M2Mφ recruitment could be correlated to tendon regeneration induced by oAECs allotransplantation. Moreover, these results demonstrate oAECs immunomodulatory role also in vivo and support novel insights into their allogeneic use underlying the resolution of tendon fibrosis.

Grafts of porcine small intestinal submucosa seeded with cultured homologous smooth muscle cells for bladder repair in dogs.

BACKGROUND: Due to numerous complications associated to gastrointestinal augmented cystoplasty, this study aimed to analyze the anatomic repair of the bladder of 10 female dogs using grafts of porcine small intestinal submucosa (SIS) seeded with cultured homologous smooth muscle cells, and compare them with the acellular SIS grafts. RESULTS: We assessed the possible side effects and complications of each type of graft by clinical examination, abdominal ultrasound and laboratory findings. Anatomic repair of neoformed bladder was assessed by histological staining for H/E and Masson's Trichrome, analyzed with a Nikon Photomicroscope connected to the system of image analysis Image J. CONCLUSIONS: We propose that SIS associated to homologous smooth cells can improve the quality of tissue repair, and consequently decrease the potential complications inherent to acellular SIS.

Cell-based therapies for tendon and ligament injuries.

Tendon and ligament injuries have proved difficult to treat effectively. Cell-based therapies offer the potential to harness the complex protein synthetic machinery of the cell to induce a regenerative response rather than fibrous scarring. This article reviews the current state of play with respect to the clinically used cell preparations for the treatment of tendon and ligaments overstrain injuries.

Cell-based therapies for equine joint disease.

Joint disease is a major cause of wastage in performance horses. Arthritis can be challenging to treat because articular cartilage has little or no capacity for repair, therapeutic options are limited and are largely targeted at ameliorating clinical signs of joint disease. Cell-based therapies have potential to overcome the intrinsic constraints to articular cartilage repair. This article focuses on cell-based therapies for treatment of equine joint disease. Results from experimental model and human clinical studies are presented along with available data from equine studies.

Commercial cell-based therapies for musculoskeletal injuries in horses.

Several cell-based therapeutic options to treat musculoskeletal injuries in horses are commercially available. The current literature supports the use of cell-based therapies to treat equine musculoskeletal injuries. Researchers continue to search for more effective cell-based therapies to provide practitioners with optimal treatment tools for musculoskeletal injuries in horses. Cell-based therapies require specialized facilities and technical competencies that might not be available or economically justifiable in many private practices. This review provides a summary of current commercially available cell-based therapeutic products for equine applications, their similarities and differences, and current objective data relating to their clinical efficacy.

In utero cell transfer between porcine littermates.

Trafficking of cells between mother and fetus during the course of normal pregnancy is well documented. Similarly, cells are known to travel between twins that share either a placenta (i.e. monozygotic) or associated chorion (i.e. monochorionic). Transferred cells are thought to be channelled via the vessels of the placenta or vascular connections established via the chorion and the long-term presence of these cells (i.e. microchimerism) can have important consequences for immune system function and reparative capacity of the host. Whether cells can be transferred between twins with separate placentas and separate chorions (i.e. no vascular connections between placentas) has not been investigated nor have the biological consequences of such a transfer. In the present study, we tested the possibility of this type of cell transfer by injecting human cord blood-derived cells into a portion of the littermates of swine and probing for human cells in the blood and tissues of unmanipulated littermates. Human cells were detected in the blood of 78% of unmanipulated littermates. Human cells were also detected in various tissues of the unmanipulated littermates, including kidney (56%), spleen (33%), thymus (11%) and heart (22%). Human cells were maintained in the blood until the piglets were sacrificed (8 months after birth), suggesting the establishment of long-term microchimerism. Our findings show that the transfer of cells between fetuses with separate placentas and separate chorions is significant and thus such twins may be subject to the same consequences of microchimerism as monozygotic or monochorionic counterparts.

Estudio de la utilidad del gel de fibrina como soporte celular en el trasplante intracerebral de c¨¦lulas madre mesenquimales / Study of the usefulness of fibrin gel as a biological scaffold for intracerebral transplantation of mesenchymal stem cells

Objetivo: Analizar el efecto de células madre del estroma de médula ósea en los traumatismos cerebrales. Material y Método: Se causó trauma cerebral por contusión, en 20 ratas Wistar adultas. A los 2 meses se transplantó CM adultas (3x106), como suspensión en suero fisiológico (volumen total de 30 µL). En 10 animales se inyectó directamente sobre la zona de lesión traumática cerebral y en otros 10 animales sobre una malla de gel de fibrina insertada en la lesión. La evolución funcional de los animales se estudió mediante los tests de mNSS y Rota-rod. Resultados: Todos los animales mostraron déficit funcional tras la lesión traumática, con mejoría significativa tras el trasplante intracerebral de CM, sin diferencias estadísticamente significativas entre ambos grupos experimentales. Conclusión: La inserción de una malla de fibrina en una lesión cerebral traumática, como soporte para la administración intralesional de CM no parece ser una técnica útil para optimizar los resultados funcionales obtenidos con este tipo de terapia celular (AU)

Objetive: To analyze the effect of intracerebral transplantation of adult stem cells obtained from bone marrow stroma in an experimental model of traumatic brain injury. Material and Method: We performed a traumatic brain contusion in 20 adult Wistar rats. At 2 months of injury, 3x106 bone marrow stromal cells as saline suspension of CM, with a total volume of 30 µL, was injected in the area of traumatic brain injury. In 10 animals the injection was made directly over the traumatic brain injury and in other 10 animals the injection was made on the mesh of fibrin gel. The functional evolution of animals was studied from injury to two months after transplantation using the mNSS test and the Rota-rod test. Results: All animals showed a clear and significant functional deficits after traumatic injury, and experienced significant improvement after intracerebral transplantation of bone marrow stromal cells, with no statistically significant differences between experimental groups. Conclusion: The insertion of a mesh of fibrin in the area of brain injury, as support for intralesional administration of CM, in an experimental model of traumatic brain injury, does not seem to be a useful technique to optimize the functional results obtained with this type of therapy cell (AU)

Refinement of in vivo surgical procedures for cardiac gene and cell transfer in rats.

In studies of gene and cell transfer for the treatment of heart disease, direct intramyocardial injection and antegrade intracoronary injection are common methods of delivering biomaterials to the heart. The authors, who carried out these surgical procedures in 377 rats, describe their methodology in detail and discuss surgical refinements that substantially reduced rat mortality. These refinements include a rigorous fluid replacement regimen, use of inhalational anesthesia instead of injectable agents, exposure of the heart without direct contact and use of a chest drainage cannula to remove air from the pleural cavity and prevent lung collapse.

Equine adipose-tissue derived mesenchymal stem cells and platelet concentrates: their association in vitro and in vivo.

Equine mesenchymal stem cells (MSC) are of particular interest both for basic research and for the therapeutic approach to musculoskeletal diseases in the horse. Their multilineage differentiation potential gives them the capability to contribute to the repair of tendon, ligament and bone damage. MSCs are also considered a promising therapeutic aid in allogeneic cell transplantation, since they show low immunogenicity and immunomodulating functions.Adipose tissue-derived adult equine stem cells (AdMSC) can be isolated, expanded in vitro and then inoculated into the damaged tissue, eventually in the presence of a biological scaffold. Here we report our preliminary experience with adipose-derived mesenchymal stem cells in allogeneic cell-therapy of tendonitis in the horse. MSCs, derived from visceral adipose tissue, were grown in the presence of autologous platelet lysate and characterized for their differentiation and growth potential. Expanded AdMSC were inoculated into the damaged tendon after their dispersion in activated platelet-rich plasma (PRP), a biological scaffold that plays an important role in maintaining cells in defect sites and contributes to tissue healing. Fourteen out of sixteen treated horses showed a functional recovery and were able to return to their normal activity.