RESUMEN
Testicular echotextural attributes are closely associated with spermatogenic development; however, precise characterisation of specific germ cell types is difficult due to tremendous germ cell heterogeneity. Recently, retinoic acid (RA) administration in neonatal mice was found to induce highly synchronised spermatogenesis as adults. A RA-treatment protocol was tested in 17 ram lambs treated with or without RA at 8 weeks of age, with scrotal ultrasonography and blood samples collected until castration 24h or 2.5 weeks later. At 8.2 weeks of age, the nuclear:seminiferous tubule (ST) area was higher in the treated compared with the control group. Serum testosterone concentrations and numerical pixel values (NPVs) of the testicular parenchyma reached a peak at 9 weeks of age in both groups of ram lambs studied. At 10.5 weeks of age, the percentage of ST cross-sections with different germ cells as the most mature germ cell type was lower and the inter-tubular heterogeneity and NPVs were also lower in the treated compared with the control animals. RA manipulation of spermatogenesis in prepubertal ram lambs may provide a suitable model for further investigation of the echotextural characteristics of specific germ cell types and critical developmental events.
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Células Germinativas/efectos de los fármacos , Testículo/efectos de los fármacos , Testosterona/sangre , Tretinoina/farmacología , Animales , Estradiol/sangre , Hormona Folículo Estimulante/sangre , Masculino , Túbulos Seminíferos/diagnóstico por imagen , Túbulos Seminíferos/efectos de los fármacos , Ovinos , Testículo/diagnóstico por imagen , UltrasonografíaRESUMEN
Cancerous cells are characterised by their ability to invade, metastasise, and induce angiogenesis. Tumour cells use various molecules that can be targeted to reverse these processes. Dasatinib, a potent Src inhibitor, has shown promising results in treating hepatocellular carcinoma (HCC) in vitro and in vivo. However, its effectiveness is limited by focal adhesion kinase (FAK) activation. Isothiocyanates, on the other hand, are phytochemicals with broad anticancer activity and FAK inhibition capabilities. This study evaluated the synergistic effect of dasatinib and phenethyl isothiocyanate (PEITC) on HCC. The combination was tested using various assays, including MTT, adhesion, scratch, Boyden chamber, chorioallantoic membrane (CAM), and yolk sac membrane (YSM) assays to evaluate the effect of the drug combination on HCC metastatic potential and angiogenesis in vitro and in vivo. The results showed that the combination inhibited the adhesion, migration, and invasion of HepG2 cells and reduced xenograft volume in the CAM assay. Additionally, the combination reduced angiogenesis in vitro, diminishing the growth of vessels in the tube formation assay. The inhibition of FAK/STAT3 signalling led to increased E-cadherin expression and reduced VEGF secretion, reducing HCC metastatic potential. Therefore, a combination of PEITC and dasatinib could be a potential therapeutic strategy for the treatment of HCC.
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Development of the scleral ossicles, a ring of bony elements within the sclera, is directed by a series of papillae that arise from placodes in the conjunctival epithelium over a 1.5-day induction period in the chicken embryo. The regular spacing of the papillae around the corneal-scleral limbus suggests that their induction may be regulated by a reaction-diffusion mechanism, similar to other epithelial appendages. Some key placode signalling molecules, including ß-catenin, are known to be expressed throughout the induction period. However, others have been studied only at certain stages or have not been successfully detected. Here we use qPCR to study the gene expression patterns of the wingless integration (WNT)/ß-catenin, bone morphogenetic protein (BMP), ectodysplasin (EDA), fibroblast growth factor (FGF) and hedgehog (HH) signalling families in discrete regions of the eye throughout the complete conjunctival placode and papillae induction period. This comprehensive analysis revealed a variable level of gene expression within specific eye regions, with some genes exhibiting high, moderate or low changes. Most genes exhibited an initial increase in gene expression, followed by a decrease or plateau as development proceeded, suggesting that some genes are important for a brief initial period whilst the sustained elevated expression level of other genes is needed for developmental progression. The timing or magnitude of these changes, and/or the overall gene expression trend differed in the temporal, nasal and/or dorsal eye regions for some, but not all genes, demonstrating that gene expression may vary across different eye regions. Temporal and nasal EDA receptor (EDAR) had the greatest number of strong correlations (r > 0.700) with other genes and ß-catenin had the greatest number of moderate correlations (r = 0.400-0.700), while EDA had the greatest range in correlation strengths. Among the strongly correlated genes, two distinct signalling modules were identified, connected by some intermediate genes. The dynamic gene expression patterns of the five signalling pathways studied here from conjunctival placode formation through to papillae development is consistent with other epithelial appendages and confirms the presence of a conserved induction and patterning signalling network. Two unique gene expression patterns and corresponding gene interaction modules suggest functionally distinct roles throughout placode development. Furthermore, spatial differences in gene expression patterns among the temporal, nasal and dorsal regions of the eye may indicate that the expression of certain genes is influenced by mechanical forces exerted throughout development. Therefore, this study identifies key placode signalling factors and their interactions, as well as some potential region-specific features of gene expression in the scleral ossicle system and provides a basis for further exploration of the spatial expression of these genes and the patterning mechanism(s) active throughout conjunctival placode and papillae formation.
Asunto(s)
Proteínas Aviares/genética , Regulación del Desarrollo de la Expresión Génica , Esclerótica/metabolismo , Vía de Señalización Wnt , Animales , Proteínas Aviares/metabolismo , Proteínas Morfogenéticas Óseas/genética , Proteínas Morfogenéticas Óseas/metabolismo , Embrión de Pollo , Conjuntiva/embriología , Conjuntiva/metabolismo , Córnea/embriología , Córnea/metabolismo , Receptor Edar/genética , Receptor Edar/metabolismo , Esclerótica/embriología , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , beta Catenina/genética , beta Catenina/metabolismoRESUMEN
The ultrasonographic image of an organ is a product of scattering and reflection of high-frequency ultrasound beams by discrete units of tissue. The number of acoustic tissue interfaces and vascularity affects the quantitative characteristics of grey-scale ultrasonographic images. This study was undertaken to examine the influences of scrotal/testicular integument and blood flow on testicular echotexture parameters in the ram. Serial ultrasonographic images were obtained during surgical castration of 7 Rideau Arcott rams aged 20-22 weeks. The first 2 sets of images were taken through the scrotum, prior to and after induction of anaesthesia. The third set was taken through the tunica vaginalis, the fourth set was obtained through the tunica albuginea, the fifth set was taken when the testicular cord and internal blood vessels were clamped, and the final set of images was recorded after allowing the blood to drain from dissected testicles (5 min). All images were then subjected to computerized image analyses and the testicles were processed for histology. The removal of the scrotal skin and tunica vaginalis both resulted in significant (P < 0.05) increments in numerical pixel values (NPVs) and pixel heterogeneity (standard deviation of pixel values) of the testicular parenchyma. There were no differences (P > 0.05) in testicular echotexture between images taken just before or after clamping the testicular cord vessels, or after draining. At all stages, NPVs were correlated (P = 0.10) to the seminiferous tubule (ST) area and the ST lumen area, except for NPVs and the ST lumen area in images obtained through the tunica albuginea (P = 0.20). We concluded that: 1) attenuation of ultrasound waves by the scrotal skin and tunica vaginalis significantly altered testicular echotexture characteristics; 2) vascular blood flow did not affect the echotextural attributes of the rams' testes; and 3) NPVs were a good indicator of ST microstructure in situ and ex vivo.
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Testículo/irrigación sanguínea , Testículo/diagnóstico por imagen , Animales , Procesamiento de Imagen Asistido por Computador , Masculino , Flujo Sanguíneo Regional/fisiología , Análisis de Regresión , Escroto/irrigación sanguínea , Escroto/diagnóstico por imagen , Ovinos , UltrasonografíaRESUMEN
The development of a skeletogenic condensation is perhaps the most critical yet considerably overlooked stage of skeletogenesis. Described in this comprehensive review are the mechanisms that facilitate skeletogenic condensation formation, growth, and maintenance to allow for overt differentiation into a skeletal element. This review discusses the current knowledge of gene regulation and characterization of skeletogenic condensations in the chicken, mouse, zebrafish, and other developmental models. We limited our scope to condensations that give rise to the bones and cartilages of the vertebrate skeleton, with a particular focus on craniofacial and limb bud regions. While many of the skeletogenic processes are similar among vertebrate lineages, differences are apparent in the site and timing of the initial epithelialâ»mesenchymal interactions as well as in whether the condensation has an osteogenic or chondrogenic fate, both within and among species. Further comparative studies are needed to clarify and broaden the existing knowledge of this intricate phenomenon.
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The objective of this study was to examine testicular ultrasonographic characteristics and endocrine profiles in prepubescent ram lambs for correlations with the age at first detection of elongated spermatids (ESt age). Bi-weekly ultrasound examinations and weekly testicular biopsies began at 10 weeks of age or at the time that testicular volume reached 15cm3, and continued until 1-2 weeks after Est's were first detected by histological examination of testicular biopsies in twenty-two spring-born Rideau Arcott×Polled Dorset lambs. Computer-assisted analysis of testicular ultrasonograms was performed using commercially available image analytical software. Blood samples were drawn before each ultrasonographic examination and were used for measurements of free triiodothyronine (fT3) and thyroxine (fT4), and follicle-stimulating hormone (FSH) concentrations. The mean (±SEM) age at first detection of ESts was 15.9±0.5 weeks. Testicular volumes recorded between 10 and 12 weeks of age correlated inversely with the ESt age (r=-0.44 to -0.50, P≤0.05). Statistically significant correlations were recorded between the ESt age and numerical pixel values of testicular parenchyma at 10 (r=-0.48, P=0.05) and 15 (r=0.52, P=0.05) weeks of age, and between the ESt age and testicular pixel heterogeneity in ram lambs aged 14.5 weeks (r=0.60, P=0.007). Lastly, circulating FSH concentrations at 10 weeks (r=-0.43, P=0.05), serum fT3 concentrations at 13 weeks (r=0.44, P=0.04) and fT4 concentrations at 11.5 weeks of age (r=0.48, P=0.03) were all correlated with the ESt age. The present results show that testicular volume has the most stable relationship with pubertal onset; however, testicular echotexture as well as circulating concentrations of FSH and free fractions of thyroid hormones at specific ages may be indicative of more intricate developmental events heralding puberty.
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Maduración Sexual/fisiología , Ovinos/fisiología , Espermatogénesis/fisiología , Testículo/diagnóstico por imagen , Ultrasonografía/veterinaria , Envejecimiento , Animales , Hormona Folículo Estimulante/sangre , Masculino , Sensibilidad y Especificidad , Ovinos/sangre , Tiroxina/sangre , Triyodotironina/sangreRESUMEN
The onset of spermatogenesis during prepubertal development is accompanied by dynamic changes in testicular microstructure. Computer-assisted analysis of scrotal ultrasonograms may allow us to track these changes in a noninvasive manner; however, the echotextural characteristics of different histomorphological variables remain unclear. Hence the objective of this study was to compare echotextural and microscopic attributes of the testis over the first wave of spermatogenesis in prepubescent ram lambs. Bi-weekly ultrasound examinations and weekly testicular biopsies were carried out in 22 ram lambs from 9.5-10 weeks of age or the attainment of 15 cm(3) in testicular volume, respectively, to the first detection of elongated spermatids (ESt). Testicular echogenicity was highly variable with age; however, after the alignment of data to the first detection of ESt, there was an initial increase followed by a decline, corresponding to the mitotic and postmitotic phases of spermatogenesis in prepubescent ram lambs. Testicular echotextural attributes (mean numerical pixel values and pixel heterogeneity) correlated with seminiferous tubule (ST) diameter, the number of degenerating cells/ST cross-section (XS), and the number of ubiquitin C-terminal hydrolase L-1 (a marker for prespermatogonia and undifferentiated spermatogonia) staining cells/ST XS during the mitotic and postmitotic phases. Additionally, in the postmitotic phase, significant correlations were recorded between the quantitative echotextural characteristics and ST cell density, nuclear:ST area and percentages of STs with different spermatogenic cells as the most mature germ cell type present. These results indicate that ram testes exhibit distinctive echotextural characteristics during the mitotic and postmitotic phases of germ cell differentiation. It is concluded that scrotal ultrasonography in conjunction with computerized image analysis holds potential as a noninvasive alternative to testicular biopsy in monitoring the reproductive status throughout different stages of testicular development.
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Histocitoquímica , Microscopía , Maduración Sexual , Ovinos/anatomía & histología , Testículo/anatomía & histología , Testículo/diagnóstico por imagen , Animales , Biopsia , Procesamiento de Imagen Asistido por Computador , Masculino , Ovinos/fisiología , Espermatogénesis , Testículo/fisiología , UltrasonografíaRESUMEN
Declining male fertility has prompted investigations into the diagnostic methods that would permit frequent, non-invasive and accurate detection of changes in testicular histomorphology and the reproductive status of individuals. Ultrasonographic (U/S) and magnetic resonance (MR) imaging both have the potential to be used in this manner as associations have previously been described between the U/S and MR image attributes and histopathological changes in testicular tissue. The present study set out to determine if correlations exist between quantitative U/S and MR image attributes and histomorphological characteristics (total and luminal seminiferous tubule, ST area, and parenchymal cell density) of the excised ram testes, and to compare relative sensitivities of the imaging techniques. The echotextural/MR (input variables) and histological parameters (output variables) were analyzed by the Pearson's product moment correlations. Significant correlations were found for all imaging modalities, with the strongest overall correlation recorded for the T2 FAST SPIN ECHO (T2FSE) MR series (between mean numerical pixel values (NPVs) and total ST area; r=-0.93, p<0.001). The greatest number of significant correlations among quantitative image characteristics and histological attributes of testicular tissue were found for the 3 PLANE LOCALIZER (3 PLANE LOC) MR series, followed by the T2FSE MR, 3D FAST-SPOILED GRADIENT ECHO (3D FSPGRE) MR, U/S (7.5 MHz) imaging, and finally T1 SPIN ECHO (T1SE) MR series. No significant correlations were recorded between the quantitative attributes of T1SE images and ST lumen area or parenchymal cell density, or between the attributes of the 3D FSPGRE images and cell density. We concluded that there existed a potential practical application for both U/S and MR image techniques, combined with computer-assisted image analysis, to monitor the changes in testicular histomorphology and male reproductive health and fertility. Scrotal U/S remains a first-line imaging technique for the assessment of male reproductive health due mainly to its versatility and lower cost.
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Imagen por Resonancia Magnética/veterinaria , Ovinos/anatomía & histología , Testículo/diagnóstico por imagen , Testículo/ultraestructura , Ultrasonografía/veterinaria , Animales , Imagen por Resonancia Magnética/métodos , Masculino , Radiografía , Ultrasonografía/métodosRESUMEN
Breeding soundness evaluation (BSE) is the primary assessment for determining the reproductive potential of male animals. This method, however, cannot be used to evaluate semen frequently or to predict future semen quality. Computerized analysis of ultrasonographic images provides information on histophysiological changes in male reproductive organs. We hypothesized that: (i) semen parameters would correlate with ultrasonographic characteristics of the distal region (cauda) of the epididymis and (ii) testicular ultrasound images and/or circulating testosterone concentration would predict future semen quality in the ram. Six adult rams underwent BSE and scrotal ultrasonography approximately 60 d apart (average duration of the spermatogenic cycle) both during the breeding (December and February) and non-breeding (June and August) seasons. An inverse correlation was found between pixel intensity (numerical pixel values) of the epididymes and percentage of sperm in semen with normal morphology (r = -0.46, P < 0.05). Pixel heterogeneity (standard deviation of pixel values) correlated negatively with percentage of sperm with normal morphology (r = -0.42, P < 0.05) and directly with percentage of spermatozoa with abnormal tails (r = 0.43, P < 0.05). Pixel heterogeneity of testicular parenchyma obtained approximately 60 d prior to semen evaluation inversely correlated with percentage of sperm with normal morphology (r = -0.73, P < 0.01) and sperm progressive motility (r = -0.76, P < 0.01), and directly with percentage of sperm with abnormal tails (r = 0.72, P < 0.01) and loose heads (r = 0.79, P < 0.01). We concluded that scrotal ultrasonography combined with computer-assisted analyses of epididymal and testicular echotexture in the ram was a valuable method for determining certain current and future semen parameters, respectively.
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Epidídimo/diagnóstico por imagen , Procesamiento de Imagen Asistido por Computador/métodos , Testículo/diagnóstico por imagen , Ultrasonografía/métodos , Animales , Humanos , Masculino , Oncología Médica/métodos , Análisis de Regresión , Semen/fisiología , Análisis de Semen , Ovinos , Recuento de Espermatozoides , Motilidad Espermática , Testículo/patologíaRESUMEN
During the last three decades, there has been remarkable progress in many aspects of ovarian biology due to advances in real-time ultrasonography, which permits non-invasive, repeated monitoring of ovarian structures in conscious and non-anaesthetised animals. This review is primarily concerned with ovarian activity, as determined by transrectal ultrasonography, and measurements of circulating concentrations of gonadotrophins and ovarian steroids during reproductive cycles in sheep. The growth of antral follicles reaching ostensibly ovulatory sizes occurs in a wave-like pattern throughout the breeding season in both prolific and non-prolific breeds of sheep. There are typically 3 or 4 waves of follicle development during the interovulatory interval. Follicular wave emergence is primarily controlled by changes in circulating concentrations of follicle-stimulating hormone (FSH) but diminished ovarian responsiveness to gonadotrophic signals may result in reduced numbers of follicular waves. In cyclic ewes, the largest ovarian follicles acquire the ability to secrete oestradiol from the day of emergence with peak oestradiol secretion occurring about the time they reach maximum diameter. The high ovulation rate in some prolific breeds may be achieved by the ovulation of follicles from the last two waves of the interovulatory interval. Prolific ewes tend to produce more but smaller corpora lutea (CL) and have lower serum concentrations of progesterone during the luteal phase of the oestrous cycle as compared to less prolific genotypes. Lastly, recent studies of the endocrine influences on ovarian function have brought into question the existence of strong follicular dominance, as seen in cattle, and provided new insights into the effects of luteal progesterone on antral follicular development in ewes.
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Ciclo Estral/fisiología , Folículo Ovárico/fisiología , Reproducción/fisiología , Ovinos/fisiología , Animales , Cuerpo Lúteo/diagnóstico por imagen , Cuerpo Lúteo/fisiología , Femenino , Masculino , Folículo Ovárico/diagnóstico por imagen , Embarazo , UltrasonografíaRESUMEN
Ovarian steroidogenesis and antral follicular development in ewes, following the treatment with medroxyprogesterone acetate (MAP) and equine chorionic gonadotrophin (eCG), are affected by the reproductive season. The objective of this study was to compare the ultrasonographic attributes of large antral follicles between cyclic (December) and seasonally anovular (June-July) ewes, after a 12-day treatment with MAP-soaked intravaginal sponges, with or without the administration of 500IU of eCG at sponge removal, and to determine whether there is a correlation between the ultrasonographic attributes of the follicular wall and serum concentrations of oestradiol. Digital images of ovulatory follicles from cyclic ewes and eCG-treated anoestrous ewes (n=34 follicles), and of anovulatory follicles attaining > or =5mm in control anoestrous ewes (n=8 follicles), were analysed using the spot and line techniques designed to determine the echotextural characteristics of the follicular antrum (central and peripheral), follicular wall and perifollicular ovarian stroma. The mean diameter of ovulatory follicles was greater (P<0.001) in cyclic than anoestrous ewes, with or without the eCG treatment. The mean pixel heterogeneity (SD of numerical pixel values) of the follicular antrum (P<0.05), as well as mean pixel intensity and heterogeneity of the peripheral antrum, follicular wall proper and perifollicular ovarian stroma (P<0.05), were consistently greater in anoestrous than cyclic ewes at the time of sponge removal and 24h after the treatment with MAP sponges or MAP/eCG. Mean oestradiol concentrations were greater (P<0.05) in cyclic compared to anoestrous ewes in both MAP- and MAP/eCG-treated animals, from 1 to 2 days after sponge withdrawal. There was a moderate negative correlation (r(2)=0.12, P<0.05; Pearson's Product Moment and r(2)=0.23, P<0.05; ANCOVA) between mean pixel heterogeneity (standard deviation of mean pixel values) of the follicular wall proper (all follicles > or =5mm in diameter) and serum concentrations of oestradiol after sponge withdrawal. Our results indicate that large antral follicles from cyclic and seasonally anovular ewes exhibit distinctive ultrasonographic characteristics. The differences in follicular echotexture appear to be related mainly to seasonal variations in ovarian follicular morphology and oestradiol production.