Maxillary sinus floor augmentation comparing bovine versus porcine bone xenografts mixed with autogenous bone graft. A split-mouth randomized controlled trial.

AIM: To compare the effectiveness of two xenografts for maxillary sinus floor augmentation in terms of clinical, radiographical, histologic, and molecular outcomes. MATERIALS AND METHODS: A split-mouth randomized clinical trial was conducted at the University of Granada. Ten consecutive patients in need of bilateral two-staged maxillary sinus floor augmentation were included. Each patient received both biomaterials (porcine bone mineral and anorganic bovine bone), which were randomly assigned for bilateral sinus augmentation. The maxillary autogenous bone scraped from the sinus access window was mixed with each xenograft at a 20:80 ratio. After a healing period of 6 months, bone biopsies were collected with a trephine during the implant placement in the regenerated area. Histologic, histomorphometrical, immunohistochemical, and molecular outcomes were analyzed. Clinical and radiographical data throughout the treatment phases were also evaluated. RESULTS: The resulting anatomic features were similar between both groups. After six months of graft consolidation, the graft resorption rates were similar between both biomaterials. The histologic, histomorphometrical, and immunohistochemical results showed no statistical differences between groups. CONCLUSION: Anorganic bovine bone and porcine bone mineral combined with maxillary autogenous cortical bone show similar biologic and radiologic features in terms of biomaterial resorption, osteoconduction, and osteogenesis when used for maxillary sinus floor augmentation.

GARP promotes the proliferation and therapeutic resistance of bone sarcoma cancer cells through the activation of TGF-ß.

Sarcomas are mesenchymal cancers with poor prognosis, representing about 20% of all solid malignancies in children, adolescents, and young adults. Radio- and chemoresistance are common features of sarcomas warranting the search for novel prognostic and predictive markers. GARP/LRRC32 is a TGF-ß-activating protein that promotes immune escape and dissemination in various cancers. However, if GARP affects the tumorigenicity and treatment resistance of sarcomas is not known. We show that GARP is expressed by human osteo-, chondro-, and undifferentiated pleomorphic sarcomas and is associated with a significantly worse clinical prognosis. Silencing of GARP in bone sarcoma cell lines blocked their proliferation and induced apoptosis. In contrast, overexpression of GARP promoted their growth in vitro and in vivo and increased their resistance to DNA damage and cell death induced by etoposide, doxorubicin, and irradiation. Our data suggest that GARP could serve as a marker with therapeutic, prognostic, and predictive value in sarcoma. We propose that targeting GARP in bone sarcomas could reduce tumour burden while simultaneously improving the efficacy of chemo- and radiotherapy.

Laminin Levels Regulate Tissue Migration and Anterior-Posterior Polarity during Egg Morphogenesis in Drosophila.

Basement membranes (BMs) are specialized extracellular matrices required for tissue organization and organ formation. We study the role of laminin and its integrin receptor in the regulation of tissue migration during Drosophila oogenesis. Egg production in Drosophila involves the collective migration of follicle cells (FCs) over the BM to shape the mature egg. We show that laminin content in the BM increases with time, whereas integrin amounts in FCs do not vary significantly. Manipulation of integrin and laminin levels reveals that a dynamic balance of integrin-laminin amounts determines the onset and speed of FC migration. Thus, the interplay of ligand-receptor levels regulates tissue migration in vivo. Laminin depletion also affects the ultrastructure and biophysical properties of the BM and results in anterior-posterior misorientation of developing follicles. Laminin emerges as a key player in the regulation of collective cell migration, tissue stiffness, and the organization of anterior-posterior polarity in Drosophila.

ECM-Regulator timp Is Required for Stem Cell Niche Organization and Cyst Production in the Drosophila Ovary.

The extracellular matrix (ECM) is a pivotal component adult tissues and of many tissue-specific stem cell niches. It provides structural support and regulates niche signaling during tissue maintenance and regeneration. In many tissues, ECM remodeling depends on the regulation of MMP (matrix metalloproteinase) activity by inhibitory TIMP (tissue inhibitors of metalloproteinases) proteins. Here, we report that the only Drosophila timp gene is required for maintaining the normal organization and function of the germline stem cell niche in adult females. timp mutant ovaries show reduced levels of both Drosophila Collagen IV α chains. In addition, tissue stiffness and the cellular organization of the ovarian niche are affected in timp mutants. Finally, loss of timp impairs the ability of the germline stem cell niche to generate new cysts. Our results demonstrating a crucial role for timp in tissue organization and gamete production thus provide a link between the regulation of ECM metabolism and tissue homeostasis.

Role of apoptosis and cell proliferation in the testicular dynamics of seasonal breeding mammals: a study in the Iberian mole, Talpa occidentalis.

Apoptosis and cell proliferation are two important cellular processes known to be involved in the normal functioning of the testis in nonseasonally breeding mammals, but there is some controversy concerning their roles in the gonads of males from seasonally breeding species. We have studied the processes of apoptosis and cell proliferation in the testes of males of the Iberian mole (Talpa occidentalis), a species showing a strict seasonal reproduction pattern. Both males and females are sexually active during the winter and completely inactive in the summer, with two transitional periods, in the autumn and the spring. Adult males from these four reproductive stages were captured, and their testes were immunohistochemically studied for the presence of apoptotic and proliferation molecular markers as well for other testicular and meiotic cell-specific markers. We found that apoptosis varies in a season-dependent manner in the testes of male moles, affecting mainly late zygotene and pachytene cells during the period of sexual inactivity, but it does not differentially affect the number of Sertoli cells. More interestingly, apoptosis is not responsible for the massive germ-cell depletion occurring during mole testis regression. In addition, a wave of spermatogonial cell proliferation appears to restore the number of spermatogonia lost during the period of testis inactivity. According to current knowledge, data from moles indicate that mammals do not form a homogeneous group regarding the mechanisms by which the cell-content dynamics are regulated in the testes of males from seasonally breeding species.

The spatio-temporal pattern of testis organogenesis in mammals - insights from the mole.

Some cellular events are crucial in testis organogenesis, including Sertoli and Leydig cell differentiation, mesonephric cell migration and testis cord formation. These processes are controlled by transcription factors, paracrine signalling and hormones. Using the mole species Talpa occidentalis as an alternative animal model, we report the expression patterns of nine genes during testis differentiation and analyse their implications in the above-mentioned cellular processes. We show that: 1) Sertoli cell differentiation occurs very early and precedes mesonephric cell migration, indicating that the latter is not needed for the endocrine cytodifferentiation of Sertoli cells; 2) the time of Leydig cell differentiation is consistent with the participation of PDGFR-alpha in promoting the migration and/or proliferation of Leydig cell precursors, and with that of WNT4 signalling in inhibiting Leydig cell differentiation and 3) the formation of the tunica albuginea involves intragonadal cell migration/movement. These results demonstrate that testicular organogenesis in the mole differs from that in the mouse in some particular aspects, thus providing evidence that the spatio-temporal pattern of testis development is not highly conserved during mammalian evolution.

SOX9 is not required for the cellular events of testicular organogenesis in XX mole ovotestes.

Mammalian sex determination is the genetic process that commits the undifferentiated bipotential gonads to develop as either testes or ovaries. The differentiation of SOX9-expressing Sertoli cells is assumed to be necessary to initiate testis development. Insectivorous moles of the genus Talpa represent a unique case of generalized true hermaphroditism, as XX female moles constitutively develop two ovotestes instead of normal ovaries. In this work, we have investigated the expression patterns of a number of genes known to play key roles in gonad organogenesis, throughout the entire process of ovotestis development in female moles. Molecular and morphological evidence are provided that these ovotestes contain primary medullary testis-like cords, Leydig cells, peritubular myoid cells, and a testis-specific vasculature, but no Sertoli cells. Our results show for the first time that SOX9 is not required for the formation of the primary testis cords, but it is necessary for the maintenance and subsequent development of these cords. In addition, the expression pattern of WNT4 in male and female moles indicates that this gene inhibits Leydig cell differentiation and, contrary to the proposed scenario in the mouse, it is not required for the colonization and survival of primordial germ cells. According to our data, mole ovotestes result from a process of PDGFRalpha-mediated mesonephric cell migration, which occurs simultaneously in both sexes. The fact that FST remains inactive during the critical stages of female gonad development, explains the lack of migration inhibition, and may be a consequence of improper WNT4 signalling in the mole.

Testis-like development of gonads in female moles. New insights on mammalian gonad organogenesis.

Moles are unique among mammals because all females of several species of genus Talpa have bilateral ovotestes (gonads with both ovarian and testicular tissue). Based on the analysis of a large sample of embryos, foetuses and infants over a 13-year period, we have studied the development of the gonads in male and female moles of the species Talpa occidentalis. Several new field and laboratory procedures were developed specifically to obtain and manage this singular material. Our results reveal that gonads of female moles develop according to a testis-like pattern, which includes cord formation and mesonephric cell migration, and begins at the same time as testis differentiation in males. The first signs of sex differentiation do not appear in males but in females. Female (but not male) gonads are regionalised with a cortex (precursor of the ovarian tissue) and a medulla (precursor of the testicular tissue). Germ cells concentrate only in the cortex, so that the medulla soon becomes sterile. Testicular tissue development is transiently retarded in females for about a week before birth, and resumes afterwards. Development of the ovarian tissue in females is considerably delayed with respect to that of testicular tissue in both males and females. The molecular characterisation of peritubular myoid cells, which are exclusive of testes, evidences the presence of testicular tissue in the gonads of female moles, which also contain Leydig cells. However, the absence of fully differentiated Sertoli cells indicates that these cells are not responsible for triggering the differentiation of such a testicular tissue. Our results are also discussed regarding the definition of Sertoli cell morphology and function, and the possible role of germ cells in the sex-reversal process. Differences observed between XX and XY gonad development in moles suggest that the mammalian testis-determining gene, SRY, has an "anti-regionalisation" role during gonadal development, at least in those mammalian species in which regionalisation of the female gonad occurs.

Abnormal sex-duct development in female moles: the role of anti-Müllerian hormone and testosterone.

We have performed a morphological, hormonal and molecular study of the development of the sex ducts in the mole Talpa occidentalis. Females develop bilateral ovotestes with a functional ovarian portion and disgenic testicular tissue. The Müllerian ducts develop normally in females and their regression is very fast in males, suggesting a powerful action of the anti-Müllerian hormone in the mole. RT-PCR demonstrated that the gene governing this hormone begins to be expressed in males coinciding with testis differentiation, and expression continues until shortly after birth. Immunohistochemical studies showed that expression occurs in the Sertoli cells of testes. No expression was detected in females. Wolffian duct development was normal in males and degenerate in prenatal females, but developmental recovery after birth gave rise to the formation of rudimentary epididymides. This event coincides in time with increasing serum testosterone levels and Leydig cell differentiation in the female gonad, thus suggesting that testosterone produced by the ovotestes is responsible for masculinisation of female moles. During postnatal development, serum testosterone concentrations decreased in males but increased in females, thus approaching the levels that adult males and females have during the non-breeding season.