As an organism is exposed to pathogens during very early development, specific defense mechanisms... more As an organism is exposed to pathogens during very early development, specific defense mechanisms must take effect. In this study, we used a germ-free zebrafish embryo model to show that osmotic stress regulates the activation of immunity and host protection in newly hatched embryos.Mechanistically, skin keratinocytes were responsible for both sensing the hyposmolarity of the aquatic environment and mediating immune effector mechanisms. This occurred through a transient potential receptor vanilloid 4/Ca2+/TGF-b–activated kinase 1/NF-kB signaling pathway. Surprisingly, the genes encoding antimicrobial effectors, which do not have the potential to cause tissue damage, are constitutively expressed during development, independently of both commensal microbes and osmotic stress. Our results reveal that osmotic stress is associated with the induction of developmental immunity in the absence of tissue damage and point out to the embryo skin as the first organ with full capacities to mount an innate immune response.
Although several studies have demonstrated the ability of some endocrine disruptive chemicals (ED... more Although several studies have demonstrated the ability of some endocrine disruptive chemicals (EDCs) to alter the physiology of zebrafish, the immune-reproductive interaction has received little attention in this species. In this study, we used a homozygous line carrying an insertion of 8 amino acids in the ligand-binding domain of the estrogen receptor 2b gene (esr2b) to further understand the role of estrogen signaling on innate immunity. Adult mutant fish showed distorted sexual ratios related with alterations in testicular morphology and supraphysiological testosterone and 17ß-estradiol (E2) levels. Immunity-wise, although esr2b mutant fish showed unaltered antibacterial responses, they were unable to mount an effective antiviral response upon viral challenge. RT-qPCR analysis demonstrated that mutant fish were able to induce the genes encoding major antiviral molecules, including Ifnphi1, Ifnphi2, Infphi3, Mxb and Mxc, and the negative feedback regulator of cytokine signaling Socs1. Notably, although esr2b mutant larvae showed a similar resistance to SVCV infection to their wild type siblings, waterborne E2 increased their viral susceptibility. Similarly, the exposure of adult wild type zebrafish to E2 also resulted in increased susceptibility to SVCV infection. Finally, the administration of recombinant Ifnphi1 hardly reversed the higher viral susceptibility of esr2b mutant zebrafish, suggesting that elevated socs1 levels impair Ifn signaling. All together, these results uncover an important role for E2 and Esr signaling in the fine-tuning of sexual hormone balance and the antiviral response of vertebrates.
Current knowledge on the sensitivity of marine fish to androgenic environmental chemicals is limi... more Current knowledge on the sensitivity of marine fish to androgenic environmental chemicals is limited, despite the growing interest in the effects of endocrine disrupting chemicals. To study in vivo the effects of testosterone (T) on the fish immune response, we used a microencapsulation implant technique, the in situ forming microparticle system, containing 1 mg T/kg body weight (T-ISM), in adult specimens of gilthead seabream (Sparus aurata L.), a species of great economic interest. We demonstrated that implants themselves (without T) have no significant effect on most of the parameters measured. In T-ISM implanted fish, T serum levels reached supraphysiological concentrations accompanied by a slight increase in 11-ketotestosterone and 17b-estradiol levels 21 days post-implantation (dpi). Liver and head-kidney samples were processed 7 and 21 dpi to assess T-ISM effect on (i) the mRNA expression of genes involved in the metabolism of steroid hormones and in the immune response, and (ii) phagocyte activities. The expression profile of cytokines, chemokines and immune receptors was altered in T-ISM implanted animals that showed an early pro-inflammatory tendency, and then, a mixed pro-/anti-inflammatory activation during longer exposure. Furthermore, the enhancement of phagocytic activity and the production of reactive oxygen species by leukocytes 21 dpi in T-ISM implanted specimens suggest fine modulation of the innate immune response by T. Taken together, these data demonstrate for the first time the feasibility of using ISM implants in an aquatic species, and provide new data on the role played by T on the immune response in fish.
Sex steroids are known to deeply alter processes other than fish reproduction, including fish gro... more Sex steroids are known to deeply alter processes other than fish reproduction, including fish growth, intermediary metabolism, osmoregulation and immunity. We have previously reported that 17β-estradiol (E(2)), the main fish estrogen, promotes the mobilization of acidophilic granulocytes from the head kidney, the bone marrow equivalent in fish, to the gonad in the bony fish gilthead seabream (Sparus aurata L.). The aim of this study was to investigate the effects of E(2) and 17α-ethinylestradiol (EE(2)), an endocrine disruptor with strong estrogenic effects commonly found in the aquatic environment, on the ability of gilthead seabream endothelial cells (ECs) to promote leukocyte infiltration. E(2) and EE(2) were seen to affect ECs in different ways. Thus, E(2) was able to increase the production of nitric oxide (NO) and up-regulate the expression of the key activation markers, interleukin-1β, CC chemokine ligand 4, interleukin-8, E-selectin and matrix metalloproteinase 9, when used alone or combined with bacterial DNA. In contrast, EE(2) failed to affect NO release and reduced the up-regulation of the above genes promoted by bacterial DNA. Moreover, we found that leukocyte adhesion to ECs was enhanced by E(2) treatment. Collectively, these results suggest that estrogens modulate fish leukocyte trafficking during an inflammatory process by activating ECs.
The role of sex steroids in the modulation of fish immune responses has received little attention... more The role of sex steroids in the modulation of fish immune responses has received little attention. Previous studies have demonstrated that 17β-estradiol (E(2)) is able to alter the response of gilthead seabream leukocytes to infectious agents. We have used suppression subtractive hybridization to identify genes upregulated by E(2) (50 ng/ml) in macrophage cultures from gilthead seabream. We isolated 393 up-regulated cDNA fragments that led to the identification of 162 candidate estrogen-responsive genes. Functional analyses revealed the presence of several enriched immune processes and molecular pathways. The E(2) up-regulation of some immune-relevant genes was further confirmed by real time RT-PCR. Bioinformatics analysis revealed the ability of E(2) to orchestrate profound alterations in the macrophage expression profile, especially immune-related processes and pathways. This is the first report on E(2)-dependent modifications of fish macrophage transcriptome and lends weight to a suggested role for estrogen in the immune system, the possible significance of which is discussed.
In mammals, estrogens regulate the immune system, either directly or indirectly via several leuko... more In mammals, estrogens regulate the immune system, either directly or indirectly via several leukocyte types through autocrine/paracrine mechanisms. In the gilthead seabream (Sparus aurata L.) gonad, an intensive remodeling process accompanied by the massive infiltration of acidophilic granulocytes (AG) is partially triggered by 17-estradiol (E 2 ). Once AG infiltrated the gonad, show impaired activities. In this study we first demonstrate that neither testicular nor head-kidney AG express any of the three estrogen receptor (ER) genes (ERa, ERb1 and ERb2) described in the gilthead seabream, while head-kidney macrophages (Mc) and lymphocytes (Ly) constitutively express ERa gene. Moreover, Mc are important in the immune-modulatory role of E 2 , as suggested by its ability to induce ERb2 gene expression and up-regulate the expression of genes coding for ERa, ERb1, pro-inflammatory cytokines, chemokines and tissue remodeling molecules. Furthermore, the soluble factors produced by E 2 -treated Mc decreased in head-kidney phagocytes, their phagocytic ability and capacity, while no effects were observed on their reactive oxygen intermediate (ROI) production or their migratory capabilities. However, the role of Ly in the regulation of AG migration and the modulation of phagocytic and ROI production activities triggered by E 2 can not be ruled out, so that further studies are necessary to clarify these issues.
Androgens can induce complete spermatogenesis in immature or prepubertal teleost fish. However, m... more Androgens can induce complete spermatogenesis in immature or prepubertal teleost fish. However, many aspects of the role of androgens in adult teleost spermatogenesis have remained elusive. Since oestrogens inhibit androgen synthesis, we used an oestrogen-induced androgen depletion model to identify androgen-dependent stages during adult zebrafish spermatogenesis. Exposure to 10 nM 17b-oestradiol (E 2 ) in vivo at least halved the mass of differentiating germ cells (from type B spermatogonia to spermatids), while type A spermatogonia accumulated. Studies on the cellular dynamics revealed that a reduction of spermatogonial proliferation together with an inhibition of their differentiation to type B spermatogonia were the basis for the oestrogen-mediated disturbance of spermatogenesis. The capacity of the zebrafish testis to produce 11-ketotestosterone as well as the expression of steroidogenesis-related genes was markedly decreased after in vivo oestrogen exposure. Moreover, the androgen-release response to recombinant zebrafish Lh was lost after oestrogen exposure. We conclude that oestrogen exposure caused a state of androgen insufficiency in adult male zebrafish. Since the downregulation of the steroidogenic system as well as the disturbance of spermatogenesis in testicular explants exposed to E 2 ex vivo was much less severe than after in vivo exposure, the main inhibitory effect appears to be exerted via feedback inhibition of gonadotropin release. This experimental set-up helped to identify spermatogonial proliferation and their differentiation as androgen targets in adult zebrafish spermatogenesis.
A great deal is known regarding the process of sex differentiation in fish. However, little is kn... more A great deal is known regarding the process of sex differentiation in fish. However, little is known about the presence of immune cells and cytokines in this process. In the gilthead seabream, both immune cells and cytokines play an important role in the tissue reorganization of the gonads during the adult reproductive cycle. We have studied, using light microscopy and immunocytochemistry, the ontogenetic development of the gilthead seabream gonads, focusing on the presence of immune cells and cytokines. We show that the testicular area is quickly differentiated and becomes functional in specimens less than a year old, while the ovarian area differentiates later and continues to develop during the first two years of life. Throughout the morphogenesis process, acidophilic granulocytes were present in the gonad. Interleukin-1b (Il1b) is produced in the testicular area in juveniles and male fish, but not in the ovarian area. Macrophage-colony stimulating factor receptor (Mcsfr) is not produced in the undifferentiated gonad and is only found once the testicular area is well developed.
Immune responses in the testis are regulated in a way that provides protection for the developing... more Immune responses in the testis are regulated in a way that provides protection for the developing male germ cells, while permitting qualitatively normal inflammatory responses and protection against infection. In addition, germ cells are potent targets for the growth factors and cytokines which regulate the reproductive process. Our study analyzes for the first time the pattern of expression of several immune-relevant genes in the gonad of a seasonal breeding teleost fish. The immune molecules analyzed include (i) inflammatory molecules, such as interleukin-1b (il1b), il6, tumor necrosis factor-a (tnfa), cyclooxygenase-2 (cox2) and the NADPH oxidase subunit p40 phox (ncf4 gene); (ii) the anti-inflammatory cytokine transforming growth factor-b1 (tgfb1) and its type 2 receptor tgfbr2; (iii) innate immune receptors, including toll-like receptor 9 (tlr9), tlr5, tlr22 and macrophage-colony stimulating factor receptor (mcsfr); (iv) lymphocyte receptors, such as the  subunit of T-cell receptor (Tcrb) and the heavy chain of immunoglobulin M (ighm); (v) the anti-bacterial molecules lysozyme (lyz), hepcidin (hamp) and complement component 3 (c3); (vi) the anti-viral molecule myxovirus (influenza) resistance protein (mx); and (vii) molecules related to leukocyte infiltration, including the CC chemokine ccl4, the CXC chemokine il8 and the leukocyte adhesion molecule E-selectin (Sele). Notably, all of them show a pattern of expression that depends on the reproductive stage of the first two reproductive cycles when the fish develop and function as males. Furthermore, we demonstrate that some of these immune-relevant molecules, such as Il1b and Mcsfr, are produced by germ cells (Il1b) and ovarian and testicular somatic cells (Mcsfr). These data suggest that, as occurs in mammals, there is a critical balance between immune molecules and that these may play an essential role in the orchestration of gametogenesis and the maintenance of gonad tissue homeostasis in fish. (A. García-Ayala).
Background: Leukocytes are found within the testis of most, if not all, mammals and are involved ... more Background: Leukocytes are found within the testis of most, if not all, mammals and are involved in immunological surveillance, physiological regulation and tissue remodelling. The testis of seasonal breeding fish undergoes a regression process. In the present study, the second reproductive cycle (RC) of the protandrous seasonal teleost fish, gilthead seabream, was investigated and the presence of leukocytes analysed. Special attention has been paid to the testicular degenerative process which is particularly active in the last stage of the second RC probably due to the immediacy of the sex change process.
The testis is a tightly controlled dynamic tissue. In mammals, there is growing evidence that est... more The testis is a tightly controlled dynamic tissue. In mammals, there is growing evidence that estrogen plays a role in the regulation of testicular functions. In teleosts, high levels of 17beta-estradiol (E 2 ) in serum correlate with the end of spermatogenesis, spawning, and the initiation of postspawning stages when spermatogonia are the main cell types in the testis. Moreover, E 2 modulates leukocyte functions in several teleost species. We hypothesized, therefore, that E 2 would induce the infiltration of acidophilic granulocytes and cause a resumption of testicular cell proliferation in spermatogenically active gilthead seabream males. Several studies of this species have reported that supraphysiological doses of E 2 are needed to induce histological and developmental changes in males. In fact, as gilthead seabream is a protandrous hermaphrodite teleost, long exposures (6-14 wk) to high doses of E 2 result in feminization of the males. Taking all this into account, we sharply increased E 2 levels during short times by i.p. injecting E 2 diluted in coconut oil as the vehicle and sampled the fish after 7, 13, and 18 days to assess the effects that E 2 had on spermatogenesis. It was observed that E 2 levels in plasma increased, while 11-ketotestosterone (11-KT) and testosterone (T) levels remained unaltered. However, 11-KT and T levels strongly increased in control fish 18 days postinjection. The most relevant result of our study was that E 2 accelerates the final events of spermatogenesis, inhibits the proliferation of spermatogonia in early stages, and induces some of the processes that usually occur during postspawning, such as the infiltration of acidophilic granulocytes and the apoptosis of primary spermatogonia. Strikingly, neither the shedding of spermatozoa nor an increase in the proliferative rate of spermatogonia stem cells was observed, probably because of the lack of other necessary stimuli, such as the increase in T levels that takes place during normal postspawning.
As an organism is exposed to pathogens during very early development, specific defense mechanisms... more As an organism is exposed to pathogens during very early development, specific defense mechanisms must take effect. In this study, we used a germ-free zebrafish embryo model to show that osmotic stress regulates the activation of immunity and host protection in newly hatched embryos.Mechanistically, skin keratinocytes were responsible for both sensing the hyposmolarity of the aquatic environment and mediating immune effector mechanisms. This occurred through a transient potential receptor vanilloid 4/Ca2+/TGF-b–activated kinase 1/NF-kB signaling pathway. Surprisingly, the genes encoding antimicrobial effectors, which do not have the potential to cause tissue damage, are constitutively expressed during development, independently of both commensal microbes and osmotic stress. Our results reveal that osmotic stress is associated with the induction of developmental immunity in the absence of tissue damage and point out to the embryo skin as the first organ with full capacities to mount an innate immune response.
Although several studies have demonstrated the ability of some endocrine disruptive chemicals (ED... more Although several studies have demonstrated the ability of some endocrine disruptive chemicals (EDCs) to alter the physiology of zebrafish, the immune-reproductive interaction has received little attention in this species. In this study, we used a homozygous line carrying an insertion of 8 amino acids in the ligand-binding domain of the estrogen receptor 2b gene (esr2b) to further understand the role of estrogen signaling on innate immunity. Adult mutant fish showed distorted sexual ratios related with alterations in testicular morphology and supraphysiological testosterone and 17ß-estradiol (E2) levels. Immunity-wise, although esr2b mutant fish showed unaltered antibacterial responses, they were unable to mount an effective antiviral response upon viral challenge. RT-qPCR analysis demonstrated that mutant fish were able to induce the genes encoding major antiviral molecules, including Ifnphi1, Ifnphi2, Infphi3, Mxb and Mxc, and the negative feedback regulator of cytokine signaling Socs1. Notably, although esr2b mutant larvae showed a similar resistance to SVCV infection to their wild type siblings, waterborne E2 increased their viral susceptibility. Similarly, the exposure of adult wild type zebrafish to E2 also resulted in increased susceptibility to SVCV infection. Finally, the administration of recombinant Ifnphi1 hardly reversed the higher viral susceptibility of esr2b mutant zebrafish, suggesting that elevated socs1 levels impair Ifn signaling. All together, these results uncover an important role for E2 and Esr signaling in the fine-tuning of sexual hormone balance and the antiviral response of vertebrates.
Current knowledge on the sensitivity of marine fish to androgenic environmental chemicals is limi... more Current knowledge on the sensitivity of marine fish to androgenic environmental chemicals is limited, despite the growing interest in the effects of endocrine disrupting chemicals. To study in vivo the effects of testosterone (T) on the fish immune response, we used a microencapsulation implant technique, the in situ forming microparticle system, containing 1 mg T/kg body weight (T-ISM), in adult specimens of gilthead seabream (Sparus aurata L.), a species of great economic interest. We demonstrated that implants themselves (without T) have no significant effect on most of the parameters measured. In T-ISM implanted fish, T serum levels reached supraphysiological concentrations accompanied by a slight increase in 11-ketotestosterone and 17b-estradiol levels 21 days post-implantation (dpi). Liver and head-kidney samples were processed 7 and 21 dpi to assess T-ISM effect on (i) the mRNA expression of genes involved in the metabolism of steroid hormones and in the immune response, and (ii) phagocyte activities. The expression profile of cytokines, chemokines and immune receptors was altered in T-ISM implanted animals that showed an early pro-inflammatory tendency, and then, a mixed pro-/anti-inflammatory activation during longer exposure. Furthermore, the enhancement of phagocytic activity and the production of reactive oxygen species by leukocytes 21 dpi in T-ISM implanted specimens suggest fine modulation of the innate immune response by T. Taken together, these data demonstrate for the first time the feasibility of using ISM implants in an aquatic species, and provide new data on the role played by T on the immune response in fish.
Sex steroids are known to deeply alter processes other than fish reproduction, including fish gro... more Sex steroids are known to deeply alter processes other than fish reproduction, including fish growth, intermediary metabolism, osmoregulation and immunity. We have previously reported that 17β-estradiol (E(2)), the main fish estrogen, promotes the mobilization of acidophilic granulocytes from the head kidney, the bone marrow equivalent in fish, to the gonad in the bony fish gilthead seabream (Sparus aurata L.). The aim of this study was to investigate the effects of E(2) and 17α-ethinylestradiol (EE(2)), an endocrine disruptor with strong estrogenic effects commonly found in the aquatic environment, on the ability of gilthead seabream endothelial cells (ECs) to promote leukocyte infiltration. E(2) and EE(2) were seen to affect ECs in different ways. Thus, E(2) was able to increase the production of nitric oxide (NO) and up-regulate the expression of the key activation markers, interleukin-1β, CC chemokine ligand 4, interleukin-8, E-selectin and matrix metalloproteinase 9, when used alone or combined with bacterial DNA. In contrast, EE(2) failed to affect NO release and reduced the up-regulation of the above genes promoted by bacterial DNA. Moreover, we found that leukocyte adhesion to ECs was enhanced by E(2) treatment. Collectively, these results suggest that estrogens modulate fish leukocyte trafficking during an inflammatory process by activating ECs.
The role of sex steroids in the modulation of fish immune responses has received little attention... more The role of sex steroids in the modulation of fish immune responses has received little attention. Previous studies have demonstrated that 17β-estradiol (E(2)) is able to alter the response of gilthead seabream leukocytes to infectious agents. We have used suppression subtractive hybridization to identify genes upregulated by E(2) (50 ng/ml) in macrophage cultures from gilthead seabream. We isolated 393 up-regulated cDNA fragments that led to the identification of 162 candidate estrogen-responsive genes. Functional analyses revealed the presence of several enriched immune processes and molecular pathways. The E(2) up-regulation of some immune-relevant genes was further confirmed by real time RT-PCR. Bioinformatics analysis revealed the ability of E(2) to orchestrate profound alterations in the macrophage expression profile, especially immune-related processes and pathways. This is the first report on E(2)-dependent modifications of fish macrophage transcriptome and lends weight to a suggested role for estrogen in the immune system, the possible significance of which is discussed.
In mammals, estrogens regulate the immune system, either directly or indirectly via several leuko... more In mammals, estrogens regulate the immune system, either directly or indirectly via several leukocyte types through autocrine/paracrine mechanisms. In the gilthead seabream (Sparus aurata L.) gonad, an intensive remodeling process accompanied by the massive infiltration of acidophilic granulocytes (AG) is partially triggered by 17-estradiol (E 2 ). Once AG infiltrated the gonad, show impaired activities. In this study we first demonstrate that neither testicular nor head-kidney AG express any of the three estrogen receptor (ER) genes (ERa, ERb1 and ERb2) described in the gilthead seabream, while head-kidney macrophages (Mc) and lymphocytes (Ly) constitutively express ERa gene. Moreover, Mc are important in the immune-modulatory role of E 2 , as suggested by its ability to induce ERb2 gene expression and up-regulate the expression of genes coding for ERa, ERb1, pro-inflammatory cytokines, chemokines and tissue remodeling molecules. Furthermore, the soluble factors produced by E 2 -treated Mc decreased in head-kidney phagocytes, their phagocytic ability and capacity, while no effects were observed on their reactive oxygen intermediate (ROI) production or their migratory capabilities. However, the role of Ly in the regulation of AG migration and the modulation of phagocytic and ROI production activities triggered by E 2 can not be ruled out, so that further studies are necessary to clarify these issues.
Androgens can induce complete spermatogenesis in immature or prepubertal teleost fish. However, m... more Androgens can induce complete spermatogenesis in immature or prepubertal teleost fish. However, many aspects of the role of androgens in adult teleost spermatogenesis have remained elusive. Since oestrogens inhibit androgen synthesis, we used an oestrogen-induced androgen depletion model to identify androgen-dependent stages during adult zebrafish spermatogenesis. Exposure to 10 nM 17b-oestradiol (E 2 ) in vivo at least halved the mass of differentiating germ cells (from type B spermatogonia to spermatids), while type A spermatogonia accumulated. Studies on the cellular dynamics revealed that a reduction of spermatogonial proliferation together with an inhibition of their differentiation to type B spermatogonia were the basis for the oestrogen-mediated disturbance of spermatogenesis. The capacity of the zebrafish testis to produce 11-ketotestosterone as well as the expression of steroidogenesis-related genes was markedly decreased after in vivo oestrogen exposure. Moreover, the androgen-release response to recombinant zebrafish Lh was lost after oestrogen exposure. We conclude that oestrogen exposure caused a state of androgen insufficiency in adult male zebrafish. Since the downregulation of the steroidogenic system as well as the disturbance of spermatogenesis in testicular explants exposed to E 2 ex vivo was much less severe than after in vivo exposure, the main inhibitory effect appears to be exerted via feedback inhibition of gonadotropin release. This experimental set-up helped to identify spermatogonial proliferation and their differentiation as androgen targets in adult zebrafish spermatogenesis.
A great deal is known regarding the process of sex differentiation in fish. However, little is kn... more A great deal is known regarding the process of sex differentiation in fish. However, little is known about the presence of immune cells and cytokines in this process. In the gilthead seabream, both immune cells and cytokines play an important role in the tissue reorganization of the gonads during the adult reproductive cycle. We have studied, using light microscopy and immunocytochemistry, the ontogenetic development of the gilthead seabream gonads, focusing on the presence of immune cells and cytokines. We show that the testicular area is quickly differentiated and becomes functional in specimens less than a year old, while the ovarian area differentiates later and continues to develop during the first two years of life. Throughout the morphogenesis process, acidophilic granulocytes were present in the gonad. Interleukin-1b (Il1b) is produced in the testicular area in juveniles and male fish, but not in the ovarian area. Macrophage-colony stimulating factor receptor (Mcsfr) is not produced in the undifferentiated gonad and is only found once the testicular area is well developed.
Immune responses in the testis are regulated in a way that provides protection for the developing... more Immune responses in the testis are regulated in a way that provides protection for the developing male germ cells, while permitting qualitatively normal inflammatory responses and protection against infection. In addition, germ cells are potent targets for the growth factors and cytokines which regulate the reproductive process. Our study analyzes for the first time the pattern of expression of several immune-relevant genes in the gonad of a seasonal breeding teleost fish. The immune molecules analyzed include (i) inflammatory molecules, such as interleukin-1b (il1b), il6, tumor necrosis factor-a (tnfa), cyclooxygenase-2 (cox2) and the NADPH oxidase subunit p40 phox (ncf4 gene); (ii) the anti-inflammatory cytokine transforming growth factor-b1 (tgfb1) and its type 2 receptor tgfbr2; (iii) innate immune receptors, including toll-like receptor 9 (tlr9), tlr5, tlr22 and macrophage-colony stimulating factor receptor (mcsfr); (iv) lymphocyte receptors, such as the  subunit of T-cell receptor (Tcrb) and the heavy chain of immunoglobulin M (ighm); (v) the anti-bacterial molecules lysozyme (lyz), hepcidin (hamp) and complement component 3 (c3); (vi) the anti-viral molecule myxovirus (influenza) resistance protein (mx); and (vii) molecules related to leukocyte infiltration, including the CC chemokine ccl4, the CXC chemokine il8 and the leukocyte adhesion molecule E-selectin (Sele). Notably, all of them show a pattern of expression that depends on the reproductive stage of the first two reproductive cycles when the fish develop and function as males. Furthermore, we demonstrate that some of these immune-relevant molecules, such as Il1b and Mcsfr, are produced by germ cells (Il1b) and ovarian and testicular somatic cells (Mcsfr). These data suggest that, as occurs in mammals, there is a critical balance between immune molecules and that these may play an essential role in the orchestration of gametogenesis and the maintenance of gonad tissue homeostasis in fish. (A. García-Ayala).
Background: Leukocytes are found within the testis of most, if not all, mammals and are involved ... more Background: Leukocytes are found within the testis of most, if not all, mammals and are involved in immunological surveillance, physiological regulation and tissue remodelling. The testis of seasonal breeding fish undergoes a regression process. In the present study, the second reproductive cycle (RC) of the protandrous seasonal teleost fish, gilthead seabream, was investigated and the presence of leukocytes analysed. Special attention has been paid to the testicular degenerative process which is particularly active in the last stage of the second RC probably due to the immediacy of the sex change process.
The testis is a tightly controlled dynamic tissue. In mammals, there is growing evidence that est... more The testis is a tightly controlled dynamic tissue. In mammals, there is growing evidence that estrogen plays a role in the regulation of testicular functions. In teleosts, high levels of 17beta-estradiol (E 2 ) in serum correlate with the end of spermatogenesis, spawning, and the initiation of postspawning stages when spermatogonia are the main cell types in the testis. Moreover, E 2 modulates leukocyte functions in several teleost species. We hypothesized, therefore, that E 2 would induce the infiltration of acidophilic granulocytes and cause a resumption of testicular cell proliferation in spermatogenically active gilthead seabream males. Several studies of this species have reported that supraphysiological doses of E 2 are needed to induce histological and developmental changes in males. In fact, as gilthead seabream is a protandrous hermaphrodite teleost, long exposures (6-14 wk) to high doses of E 2 result in feminization of the males. Taking all this into account, we sharply increased E 2 levels during short times by i.p. injecting E 2 diluted in coconut oil as the vehicle and sampled the fish after 7, 13, and 18 days to assess the effects that E 2 had on spermatogenesis. It was observed that E 2 levels in plasma increased, while 11-ketotestosterone (11-KT) and testosterone (T) levels remained unaltered. However, 11-KT and T levels strongly increased in control fish 18 days postinjection. The most relevant result of our study was that E 2 accelerates the final events of spermatogenesis, inhibits the proliferation of spermatogonia in early stages, and induces some of the processes that usually occur during postspawning, such as the infiltration of acidophilic granulocytes and the apoptosis of primary spermatogonia. Strikingly, neither the shedding of spermatozoa nor an increase in the proliferative rate of spermatogonia stem cells was observed, probably because of the lack of other necessary stimuli, such as the increase in T levels that takes place during normal postspawning.
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Papers by S. Liarte
immunity in the absence of tissue damage and point out to the embryo skin as the first organ with full capacities to mount an
innate immune response.
immunity in the absence of tissue damage and point out to the embryo skin as the first organ with full capacities to mount an
innate immune response.