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  • 1.
    Alvarez-Rodriguez, Manuel
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Vicente Carrillo, Alejandro
    Linköpings universitet, Medicinska fakulteten. Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Evidensia Valla Djursjukhus Linkoping, Linkoping, Sweden.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Hyaluronan improves neither the long-term storage nor the cryosurvival of liquid-stored CD44-bearing Al boar spermatozoa2018Ingår i: Journal of reproduction and development, ISSN 0916-8818, E-ISSN 1348-4400, Vol. 64, nr 4, s. 351-360Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Hyaluronan (hyaluronic acid, HA) apparently improves sperm survival in vitro and in vivo (oviduct), maintaining sperm motility and inducing capacitation, but not acrosome exocytosis, either by direct action as a macromolecule or via CD44 membrane receptors. This study explored ejaculated, liquid-extended pig spermatozoa to ascertain (i) the presence (Western blotting) and specific location (immunocytochemistry) of the CD44 receptor, using a specific monoclonal commercial antibody; (ii) whether the CD44 receptor changed location when exposed to bicarbonate, a capacitating trigger, in vitro; and (iii) whether the addition of HA, of molecular size comparable to that produced in the oviduct sperm reservoir (0.0625 to 2.0 mg/ml; 0 HA: control), to semen extenders would improve sperm liquid storage in vitro or cryosurvival post freezing. Variables tested were sperm velocity and progressive motility (Qualisperm (TM)), sperm viability and acrosome status, membrane integrity and early destabilization, mitochondrial activation, and superoxide production (flow cytometry). The CD44 receptor presence in ejaculated, liquid-stored AI boar spermatozoa, as confirmed by a porcine-specific monoclonal antibody, maintained its membrane location under in vitro capacitation-inducing conditions. HA exposure to 24-, 48-, or 72-h liquid-stored (17-20 degrees C) spermatozoa lowered sperm velocity in membrane-intact spermatozoa, but increased mitochondrial superoxide production. Finally, HA addition during cooling did not improve cryosurvival but did increase mitochondrial activation and membrane destabilization in surviving cells. These results confirm the existence of a CD44 receptor in pig spermatozoa, but the usefulness of adding HA for long-term storage or cryopreservation of liquid-stored, extended boar semen remains in question, thereby warranting further non-empirical analyses of HA-sperm membrane interactions.

  • 2.
    Alvarez-Rodriguez, Manuel
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Vicente-Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Exogenous Individual Lecithin-Phospholipids (Phosphatidylcholine and Phosphatidylglycerol) Cannot Prevent the Oxidative Stress Imposed by Cryopreservation of Boar Sperm.2017Ingår i: Journal of veterinary medicine and surgery, ISSN 2574-2868, Vol. 1, nr 1Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Objective: Despite the use of high proportions of the chemically undefined lipoprotein/phospholipid-rich egg-yolk in extenders, boar sperm are highly sensitive to cooling, which induces ROS generation and disrupts the plasma membrane.

    Here, we studied whether replacement of hen egg-yolk by commercially defined lecithin phospholipids, derived from egg (LPGE: phosphatidyl glycerol, LPCE: phosphatidyl choline) or soybean (LPCS: phosphatidyl choline), could individually ameliorate such oxidative effects during cryopreservation of ejaculated (sperm rich fraction, SRF) or of cauda-epididymal sperm, retrieved post-mortem from the same males.

    Methods: A conventional extender (lactose buffer, with 20% egg-yolk, 0.5% OEP and 3% glycerol) was used as control. Cryodamage was assessed as loss of sperm motility, membrane and acrosome intactness, early membrane destabilization changes, mitochondrial potential, superoxide and ROS production, to finally determine lipid peroxidation (LPO) using specific probes.

    Results and conclusion: In general, the exogenous phospholipids assayed were unable of maintaining neither sperm motility nor viability post-thaw compared to controls, owing to increased ROS production and lipid peroxidation. In our study, mitochondrial superoxide production resulted in very high levels for all groups, whereas both ROS production and lipid peroxidation were reduced in the control group, containing emulsified hen egg yolk. Further studies using various dosage and combination of LPCS should be followed for their eventual protective effect.

    Keywords: Cryodamage; Sperm; Boar; Mitochondrial activation; Mitochondrial superoxide; ROS production; Lipid peroxidation

  • 3.
    Atikuzzaman, Mohammad
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Alvarez-Rodriguez, Manuel
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Carrillo, Alejandro Vicente
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Johnsson, Martin
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Wright, Dominic
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Conserved gene expression in sperm reservoirs between birds and mammals in response to mating.2017Ingår i: BMC Genomics, ISSN 1471-2164, E-ISSN 1471-2164, Vol. 18, nr 1Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    BACKGROUND: Spermatozoa are stored in the oviductal functional sperm reservoir in animals with internal fertilization, including zoologically distant classes such as pigs or poultry. They are held fertile in the reservoir for times ranging from a couple of days (in pigs), to several weeks (in chickens), before they are gradually released to fertilize the newly ovulated eggs. It is currently unknown whether females from these species share conserved mechanisms to tolerate such a lengthy presence of immunologically-foreign spermatozoa. Therefore, global gene expression was assessed using cDNA microarrays on tissue collected from the avian utero-vaginal junction (UVJ), and the porcine utero-tubal junction (UTJ) to determine expression changes after mating (entire semen deposition) or in vivo cloacal/cervical infusion of sperm-free seminal fluid (SF)/seminal plasma (SP).

    RESULTS: In chickens, mating changed the expression of 303 genes and SF-infusion changed the expression of 931 genes, as compared to controls, with 68 genes being common to both treatments. In pigs, mating or SP-infusion changed the expressions of 1,722 and 1,148 genes, respectively, as compared to controls, while 592 genes were common to both treatments. The differentially expressed genes were significantly enriched for GO categories related to immune system functions (35.72-fold enrichment). The top 200 differentially expressed genes of each treatment in each animal class were analysed for gene ontology. In both pig and chicken, an excess of genes affecting local immune defence were activated, though frequently these were down-regulated. Similar genes were found in both the chicken and pig, either involved in pH-regulation (SLC16A2, SLC4A9, SLC13A1, SLC35F1, ATP8B3, ATP13A3) or immune-modulation (IFIT5, IFI16, MMP27, ADAMTS3, MMP3, MMP12).

    CONCLUSION: Despite being phylogenetically distant, chicken and pig appear to share some gene functions for the preservation of viable spermatozoa in the female reservoirs.

  • 4.
    Atikuzzaman, Mohammad
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Alvarez-Rodriguez, Manuel
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Vicente Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Johnsson, Martin
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Wright, Dominic
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Correction: Conserved gene expression in sperm reservoirs between birds and mammals in response to mating (vol 18, 98, 2017)2017Ingår i: BMC Genomics, ISSN 1471-2164, E-ISSN 1471-2164, Vol. 18, artikel-id 563Artikel i tidskrift (Övrigt vetenskapligt)
    Abstract [en]

    n/a

  • 5.
    Barranco, Isabel
    et al.
    University of Murcia, Murcia, Spain.
    Perez-Patiño, Cristina
    University of Murcia, Murcia, Spain.
    Tvarijonaviciute, Asta
    University of Murcia, Murcia, Spain.
    Parrilla, Inmaculada
    University of Murcia, Murcia, Spain.
    Vicente-Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Alvarez-Rodriguez, Manuel
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Ceron, Jose J
    University of Murcia, Murcia, Spain.
    Martinez, Emilio A
    University of Murcia, Murcia, Spain.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Roca, Jordi
    University of Murcia, Murcia, Spain.
    Active paraoxonase 1 is synthesised throughout the internal boar genital organs.2017Ingår i: Reproduction (Cambridge, England), ISSN 1741-7899, Vol. 154, nr 3, s. 237-243Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The paraoxonase type 1 (PON1) is an enzyme with antioxidant properties recently identified in the seminal plasma (SP) of several species, including the porcine. The aims of the present study were to (1) describe the immunohistochemical localisation of PON1 in the genital organs of fertile boars and (2) evaluate the relationship among PON1 activity and high-density lipoprotein cholesterol (HDL-C) concentration in fluids of the boar genital organs. Immunohistochemical analysis demonstrated that PON1 was present in testis (specifically in Leydig cells, blood vessels, spermatogonia and elongated spermatids), epididymis (specifically in the cytoplasm of the principal epithelial cells, luminal secretion and in the surrounding smooth muscle) and the lining epithelia of the accessory sexual glands (cytoplasmic location in the prostate and membranous in the seminal vesicle and bulbourethral glands). The Western blotting analysis confirmed the presence of PON1 in all boar genital organs, showing in all of them a band of 51 kDa and an extra band of 45 kDa only in seminal vesicles. PON1 showed higher activity levels in epididymal fluid than those in SP of the entire ejaculate or of specific ejaculate portions. A highly positive relationship between PON1 activity and HDL-C concentration was found in all genital fluids. In sum, all boar genital organs contributing to sperm-accompanying fluid/s were able to express PON1, whose activity in these genital fluids is highly dependent on the variable HDL-C concentration present.

  • 6.
    Barranco, Isabel
    et al.
    University of Murcia, Spain.
    Roca, Jordi
    University of Murcia, Spain.
    Tvarijonaviciute, Asta
    University of Murcia, Spain.
    Rubér, Marie
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Vicente Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Hälsouniversitetet.
    Atikuzzaman, Mohammad
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Ceron, Jose J.
    University of Murcia, Spain.
    Martinez, Emilio A.
    University of Murcia, Spain.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Measurement of Activity and Concentration of Paraoxonase 1 (PON-1) in Seminal Plasma and Identification of PON-2 in the Sperm of Boar Ejaculates2015Ingår i: Molecular Reproduction and Development, ISSN 1040-452X, E-ISSN 1098-2795, Vol. 82, nr 1, s. 58-65Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    This study revealed and characterised the presence of the antioxidant enzymes paraoxonase (PON) type 1 (PON-1, extracellular) and type 2 (PON-2, intracellular) in boar semen. To evaluate PON-1, an entire ejaculate from each of ten boars was collected and the seminal plasma was harvested after double centrifugation (1,500g for 10min). Seminal plasma was analysed for concentration as well as enzymatic activity of PON-1 and total cholesterol levels. Seminal-plasma PON-1 concentration ranged from 0.961 to 1.670ng/ml while its enzymatic activity ranged from 0.056 to 0.400 IU/ml, which represent individual variance. Seminal-plasma PON-1 concentration and enzymatic activity were negatively correlated (r=-0.763; Pless than0.01). The activity of seminal-plasma PON-1 negatively correlated with ejaculate volume (r=-0.726, Pless than0.05), but positively correlated with sperm concentration (r=0.654, Pless than0.05). Total seminal-plasma cholesterol concentration positively correlated with PON-1 activity (r=0.773; Pless than0.01), but negatively correlated with PON-1 concentration (r=-0.709; Pless than0.05). The presence of intracellular PON-2 was determined via immunocytochemistry in spermatozoa derived from artificial insemination. PON-2 localised to the post-acrosomal area of the sperm head and principal piece of the tail in membrane-intact spermatozoa. In summary, PON is present in boar semen, with PON-1 at low levels in seminal plasma and PON-2 within the spermatozoa. Further studies are needed to characterise the relationship between antioxidant PONs with sperm and other seminal-plasma parameters. Mol. Reprod. Dev. 82: 58-65, 2015. (c) 2014 Wiley Periodicals, Inc.

  • 7.
    Barranco, Isabel
    et al.
    University of Murcia, Spain.
    Tvarijonaviciute, Asta
    University of Murcia, Spain.
    Perez-Patino, Cristina
    University of Murcia, Spain.
    Vicente Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Parrilla, Inmaculada
    University of Murcia, Spain.
    Ceron, Jose J.
    University of Murcia, Spain.
    Martinez, Emilio A.
    University of Murcia, Spain.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Roca, Jordi
    University of Murcia, Spain.
    Glutathione Peroxidase 5 Is Expressed by the Entire Pig Male Genital Tract and Once in the Seminal Plasma Contributes to Sperm Survival and In Vivo Fertility2016Ingår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 11, nr 9, artikel-id e0162958Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Glutathione peroxidase-5 (GPX5) is an H2O2-scavenging enzyme identified in boar seminal plasma (SP). This study attempted to clarify its origin and role on sperm survival and fertility after artificial insemination (AI). GPX5 was expressed (Western blot and immunocytochemistry using a rabbit primary polyclonal antibody) in testes, epididymis and accessory sex glands (6 boars). SP-GPX5 concentration differed among boars (11 boars, P amp;lt; 0.001), among ejaculates within boar (44 ejaculates, P amp;lt; 0.001) and among portions within ejaculate (15 ejaculates). The first 10 mL of the spermrich fraction (SRF, sperm-peak portion) had a significantly lower concentration (8.87 +/- 0.78 ng/mL) than the rest of the SRF and the post-SRF (11.66 +/- 0.79 and 12.37 +/- 0.79 ng/mL, respectively, P amp;lt; 0.005). Spermmotility of liquid-stored semen AI-doses (n = 44, at 15-17 degrees C during 72h) declined faster in AI-doses with low concentrations of SP-GPX5 compared to those with high-levels. Boars (n = 11) with high SP-GPX5 showed higher farrowing rates and litter sizes than those with low SP-GPX5 (a total of 5,275 inseminated sows). In sum, GPX5 is widely expressed in the boar genital tract and its variable presence in SP shows a positive relationship with sperm quality and fertility outcomes of liquid-stored semen AI-doses.

  • 8.
    Gonzalez-Arto, Marta
    et al.
    University of Zaragoza, Spain.
    Vicente Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Martinez-Pastor, Felipe
    University of Leon, Spain.
    Fernandez-Alegre, Estela
    University of Leon, Spain.
    Roca, Jordi
    University of Murcia, Spain.
    Miro, Jordi
    University of Barcelona, Spain.
    Rigau, Teresa
    University of Barcelona, Spain.
    Rodriguez-Gil, Joan E.
    University of Barcelona, Spain.
    Perez-Pe, Rosaura
    University of Zaragoza, Spain.
    Muino-Blanco, Teresa
    University of Zaragoza, Spain.
    Cebrian-Perez, Jose A.
    University of Zaragoza, Spain.
    Casao, Adriana
    University of Zaragoza, Spain.
    Melatonin receptors MT1 and MT2 are expressed in spermatozoa from several seasonal and nonseasonal breeder species2016Ingår i: Theriogenology, ISSN 0093-691X, E-ISSN 1879-3231, Vol. 86, nr 8, s. 1958-1968Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Melatonin is a ubiquitous and multipurpose molecule, and one of its roles is to regulate reproduction in some seasonal mammals. Our group has previously reported the variation in the melatonin levels in ram seminal plasma along the year and identified MT1 and MT2 receptors in ram spermatozoa. The objective of this study was to elucidate whether the presence of melatonin receptors (MT1 and MT2) in the sperm plasma membrane, and melatonin in the seminal plasma is related to seasonal breeding. For this purpose, the presence of melatonin receptors and tseasonal breeder (epididymal spermatozoa); bull as a conventional nonseasonhe levels of melatonin in seminal plasma have been examined in several species: donkey and stallion as long-day breeders; red deer as a wild, short-day, highly al breeder; boar as a seasonal breeder, under management techniques; and dog as possible a seasonal breeder not regulated by melatonin. We have detected measurable levels of melatonin in the seminal plasma of all ejaculated semen samples (from donkey, stallion, boar, bull, and dog). Also, and for the first time, we have demonstrated the presence of MT1 and MT2 melatonin receptors in the spermatozoa of all these species, regardless their type of reproduction or sperm source (ejaculated or epididymal), using indirect immunofluorescence techniques and Western blotting. Our findings suggest that melatonin and melatonin receptors may be universally distributed in the reproductive system of mammals and that the sperm melatonin receptors cells may not be necessarily related with seasonal reproduction. Furthermore, the presence of MT1 at the cytoplasmic droplet in immature ejaculated stallion spermatozoa found in one sample and epididymal red deer spermatozoa suggests that melatonin may be involved in specific functions during spermatogenesis and sperm maturation, like protecting spermatozoa from oxidative damage, this activity being mediated through these receptors. (C) 2016 Elsevier Inc. All rights reserved.

  • 9.
    Iso-Touru, Terhi
    et al.
    Nat Resources Inst Finland Luke, Finland.
    Wurmser, Christine
    Tech Univ Munich, Germany.
    Venhoranta, Heli
    Univ Helsinki, Finland.
    Hiltpold, Maya
    Swiss Fed Inst Technol, Switzerland.
    Savolainen, Tujia
    Univ Helsinki, Finland.
    Sironen, Anu
    Nat Resources Inst Finland Luke, Finland.
    Fischer, Konrad
    Tech Univ Munich, Germany.
    Flisikowski, Krzysztof
    Tech Univ Munich, Germany.
    Fries, Ruedi
    Tech Univ Munich, Germany.
    Vicente Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin. Linköpings universitet, Medicinska fakulteten.
    Alvarez-Rodriguez, Manuel
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Nagy, Szabolcs
    Univ Pannonia, Hungary.
    Mutikainen, Mervi
    Nat Resources Inst Finland Luke, Finland.
    Peippo, Jaana
    Nat Resources Inst Finland Luke, Finland.
    Taponen, Juhani
    Univ Helsinki, Finland.
    Sahana, Goutam
    Aarhus Univ, Denmark.
    Guldbrandtsen, Bernt
    Aarhus Univ, Denmark.
    Simonen, Henri
    VikingGenet, Finland.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Andersson, Magnus
    Univ Helsinki, Finland.
    Pausch, Hubert
    Swiss Fed Inst Technol, Switzerland.
    A splice donor variant in CCDC189 is associated with asthenospermia in Nordic Red dairy cattle2019Ingår i: BMC Genomics, ISSN 1471-2164, E-ISSN 1471-2164, Vol. 20, nr 1, artikel-id 286Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Background

    Cattle populations are highly amenable to the genetic mapping of male reproductive traits because longitudinal data on ejaculate quality and dense microarray-derived genotypes are available for thousands of artificial insemination bulls. Two young Nordic Red bulls delivered sperm with low progressive motility (i.e., asthenospermia) during a semen collection period of more than four months. The bulls were related through a common ancestor on both their paternal and maternal ancestry. Thus, a recessive mode of inheritance of asthenospermia was suspected.

    Results

    Both bulls were genotyped at 54,001 SNPs using the Illumina BovineSNP50 Bead chip. A scan for autozygosity revealed that they were identical by descent for a 2.98 Mb segment located on bovine chromosome 25. This haplotype was not found in the homozygous state in 8557 fertile bulls although five homozygous haplotype carriers were expected (P = 0.018). Whole genome-sequencing uncovered that both asthenospermic bulls were homozygous for a mutation that disrupts a canonical 5′ splice donor site of CCDC189 encoding the coiled-coil domain containing protein 189. Transcription analysis showed that the derived allele activates a cryptic splice site resulting in a frameshift and premature termination of translation. The mutated CCDC189 protein is truncated by more than 40%, thus lacking the flagellar C1a complex subunit C1a-32 that is supposed to modulate the physiological movement of the sperm flagella. The mutant allele occurs at a frequency of 2.5% in Nordic Red cattle.

    Conclusions

    Our study in cattle uncovered that CCDC189 is required for physiological movement of sperm flagella thus enabling active progression of spermatozoa and fertilization. A direct gene test may be implemented to monitor the asthenospermia-associated allele and prevent the birth of homozygous bulls that are infertile. Our results have been integrated in the Online Mendelian Inheritance in Animals (OMIA) database (https://omia.org/OMIA002167/9913/).

  • 10.
    Rodriguez-Martinez, Heriberto
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Tienthai, P.
    Chulalongkorn University, Thailand.
    Atikuzzaman, Mohammad
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Vicente Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Rubér, Marie
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Alvarez-Rodriguez, Manuel
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    The ubiquitous hyaluronan: Functionally implicated in the oviduct?2016Ingår i: Theriogenology, ISSN 0093-691X, E-ISSN 1879-3231, Vol. 86, nr 1, s. 182-186Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    Hyaluronan (hyaluronic acid) is a simple, nonantigenic, nonsulfated glycosaminoglycan present everywhere in the extracellular compartments of the body. Noteworthy, it is highly conserved phylogenetically, from sauropsida to mammals; and plays a plethora of roles from embryonic/fetal development to adult physiological and pathological events, including tumor development. In reproduction, hyaluronan has proven related to initial events as sperm survival, buildup of the sperm reservoir in the oviduct, regulation of sperm capacitation, and prefertilization to later participate in embryo, fetal, and placental development. Synthesis, binding (via the CD44 membrane receptor), and degradation of hyaluronan occur in male and female genital organs, the oviduct being no exception. This review discusses our current knowledge on roles of this ubiquitous glycosaminoglycan on the survival of immunologically foreign spermatozoa in the pig oviduct, a relevant event for fertility. During preovulatory storage in the functional tubal sperm reservoir, spermatozoa are entrapped in a mucus-like tubal fluid. This fluid contains fluctuating levels of hyaluronan, which is synthesized by the lining epithelium by hyaluronan synthase 3. Both hyaluronan and its CD44 receptor are particularly evident in the deep mucosal furrows of the sperm reservoir, in which most spermatozoa are embedded in; kept alive, uncapacitated but also undetected by the immune system of the female. Hyaluronan is also present in the seminal plasma, and evidence points toward an involvement of hyaluronan and its receptor in the local (tubal and possibly uterine) production of antiinflammatory cytokines, such as interleulcin-10, pertaining maternal immune tolerance of these foreign cells. (C) 2015 Elsevier Inc. All rights reserved.

  • 11.
    Sanchez Centellas, Daniel
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Medicinska fakulteten.
    Gudlur, Sushanth
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Medicinska fakulteten. Nanyang Technology University, Singapore.
    Vicente Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Ramström, Sofia
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Medicinska fakulteten.
    Lindahl, Tomas
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Medicinska fakulteten. Region Östergötland, Diagnostikcentrum, Klinisk kemi.
    A cluster of aspartic residues in the extracellular loop II of PAR 4 is important for thrombin interaction and activation of platelets2017Ingår i: Thrombosis Research, ISSN 0049-3848, E-ISSN 1879-2472, Vol. 154, s. 98-106Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Thrombin activates platelets via proteolytic cleavage of protease-activated receptors (PARs) 1 and 4. The two PARs have distinct but complementary roles. The mechanisms responsible for PAR1 activation by thrombin have been extensively studied. However, much less is known regarding thrombin activation of PAR4, especially the potential involvement of regions of PAR4 other than the N-terminal, which is bound to the catalytic site of thrombin. We have studied PAR4 in S. cerevisiae strainMMY12, an expression system in which the GPCR receptors are connected to a Lac Z reporter gene resulting in increased beta-galactosidase activity. This approach was used to assess PAR4 mutants to evaluate the contribution of different aspartic residues in facilitating PAR4 activation. Furthermore, peptides mimicking parts of the PAR4 N-terminal and the second extracellular loop (ECLII) were tested for their ability to inhibit platelet activation by thrombin. Binding of these peptides to gamma-thrombin was studied by monitoring the decrease in tryptophan fluorescence intensity of thrombin. We conclude that not only the N-terminal but also the electronegative aspartic residues D224, D230 and D235 (located in ECLII) are be important for PAR4 binding to thrombin. We further suggest that they play a role for the tethered ligand binding to the receptor, as mutations also affected activation in response to a PAR4-activating peptide mimicking the new N-terminal formed after cleavage. This agrees with previous results on PAR1 and thrombin binding. We suggest that the ECLII of PAR4 could be a potential target for antithrombotic drug development. (C) 2017 Elsevier Ltd. All rights reserved.

  • 12.
    Vicente Carrillo, Alejandro
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Sperm Membrane Channels, Receptors and Kinematics: Using boar spermatozoa for drug toxicity screening2016Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Internal fertilization usually implies that a spermatozoon, with intact attributes for zygote formation, passes all hurdles during its transport through the female genitalia and reaches the oocyte. During this journey, millions to billions of other spermatozoa perish. Spermatozoa are highly differentiated motile cells without synthetic capabilities. They generate energy via glycolysis and oxidative phosphorylation to sustain motility and to maintain the stability and functionality of their plasma membrane. In vivo, they spend their short lifespan bathing in female genital tract fluids of different origins, or are in vitro exposed to defined media during diverse sperm handling i.e. extension, cryopreservation, in vitro fertilization, etc. Being excitable cells, spermatozoa respond in vivo to various stimuli during pre-fertilization (capacitation, hyperactivation, oocyte location) and fertilization (acrosome reaction, interaction with the oocyte) events, mediated via diverse membrane ion-conducting channels and ligand-gated receptors. The present Thesis has mapped the presence and reactivity (sperm intactness and kinematics) of selected receptors, water and ion channels in ejaculated boar spermatozoa. The final aim was to find a relevant alternative cell type for in vitro bioassays that could ease the early scrutiny of candidate drugs as well as decreasing our needs for experimental animals according to the 3R principles. Spermatozoa are often extended, cooled and thawed to warrant their availability as fertile gametes for breeding or in vitro testing. Such manipulations stress the cells via osmotic variations and hence spermatozoa need to maintain membrane intactness by controlling the exchange of water and the common cryoprotectant glycerol, via aquaporins (AQPs). Both AQPs-7 and -9 were studied for membrane domain changes in cauda- and ejaculated spermatozoa (un-processed, extended, chilled or frozen-thawed). While AQP-9 maintained location through source and handling, thawing of ejaculated spermatozoa clearly relocated the labelling of AQP-7, thus appearing as a relevant marker for non-empirical studies of sperm cryopreservation. Alongside water, spermatozoa interact with calcium (Ca2+) via the main Ca2+ sperm channel CatSper. Increments in intracellular Ca2+ initiate motility hyperactivation and the acrosome reaction. The four subunits of the CatSper channel were present in boar spermatozoa, mediating changes in sperm motility under in vitro capacitation-inducing conditions (increased extracellular Ca2+ availability and bicarbonate) or challenge by the CatSper antagonists mibefradil and NNC 55-0396. Uterine and oviduct fluids are richest in endogenous opioids as β-endorphins during mating and ovulation. Both μ- and δ- opioid receptors were present in boar spermatozoa modulating sperm motility, as in vitro challenge with known agonists (μ: morphine; δ: DPDPE and κ: U 50488) and antagonists (μ: naloxone; δ: naltrindole and κ: nor-binaltrorphimine) showed that the μ-opioid receptor maintained or increased motility while the δ-opioid receptor mediated decreased motility over time. Finally, boar spermatozoa depicted dose-response effects on sperm kinematics and mitochondrial potential following in vitro challenge with 130 pharmacological drugs and toxic compounds as well as with eight known mito-toxic compounds. In conclusion, boar spermatozoa expressing functional water (AQPs-7 and -9) and ion (CatSper 1-4) channels as well as μ- and δ-opioid receptors are able to adapt to stressful environmental variations, capacitation and pharmacological compounds and drug components. Ejaculated sperm suspensions are easily and painlessly obtained from breeding boars, and are suitable biosensors for in vitro drug-induced testing, complying with the 3R principles of reduction and replacement of experimental animals, during early toxicology screening.

    Delarbeten
    1. Membrane Stress During Thawing Elicits Redistribution of Aquaporin 7 But Not of Aquaporin 9 in Boar Spermatozoa
    Öppna denna publikation i ny flik eller fönster >>Membrane Stress During Thawing Elicits Redistribution of Aquaporin 7 But Not of Aquaporin 9 in Boar Spermatozoa
    2016 (Engelska)Ingår i: Reproduction in domestic animals, ISSN 0936-6768, E-ISSN 1439-0531, Vol. 51, nr 5, s. 665-679Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Freezing of boar spermatozoa includes the cryoprotectant glycerol, but renders low cryosurvival, owing to major changes in osmolarity during freezing/thawing. We hypothesize that aquaporins (AQPs) 7 and 9 adapt their membrane domain location to these osmotic challenges, thus maintaining sperm homeostasis. Western blotting (WB) and immunocytochemistry (ICC) at light and electron microscope levels with several commercial primary antibodies and protocols explored AQP location on cauda epididymal and ejaculated spermatozoa (from different fractions of the ejaculate), unprocessed, extended, chilled and frozen-thawed. Although differences in WB and ICC labelling were seen among antibodies, AQP-7 was conspicuously located in the entire tail and cytoplasmic droplet in caudal spermatozoa, being restricted to the mid-piece and principal piece domains in ejaculated spermatozoa. AQP-9 was mainly localized in the sperm head in both caudal and ejaculated spermatozoa. While unaffected by chilling (+5°C), freezing and thawing of ejaculated spermatozoa clearly relocated the head labelling of AQP-7, but not that of AQP-9. In vitro mimicking of cell membrane expansion during quick thawing maintained the localization of AQP-9 but relocated AQP-7 towards the acrosome. AQP-7, but not AQP-9, appears as a relevant marker for non-empirical studies of sperm handling.

    Ort, förlag, år, upplaga, sidor
    Blackwell Verlag, 2016
    Nationell ämneskategori
    Utvecklingsbiologi
    Identifikatorer
    urn:nbn:se:liu:diva-130228 (URN)10.1111/rda.12728 (DOI)000388334100005 ()27405395 (PubMedID)
    Anmärkning

    Funding agencies: Swedish Research Council VR, Stockholm [521-2011-6553]; Research Council FORMAS, Stockholm [221-2011-512]; FORSS (Forskningsradet i Sydostra Sverige), Sweden [473121]

    Tillgänglig från: 2016-07-19 Skapad: 2016-07-19 Senast uppdaterad: 2018-03-23Bibliografiskt granskad
    2. The CatSper channel modulates boar sperm motility during capacitation.
    Öppna denna publikation i ny flik eller fönster >>The CatSper channel modulates boar sperm motility during capacitation.
    2017 (Engelska)Ingår i: Reproductive biology, ISSN 2300-732X, Vol. 17, nr 1, s. 69-78Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    The cation channel of sperm (CatSper) comprises four transmembrane subunits specifically expressed in human, equine, murine and ovine spermatozoa, apparently implicated in capacitation, hyperactivation and acrosome exocytosis. Western blotting and immunocytochemistry showed hereby that CatSper subunits are also present in boar spermatozoa, primarily over the sperm neck, tail and cytoplasmic droplets; albeit CatSper -1 presented in addition some distribution over the membrane of the acrosome and CatSper -2 and -4 over the membrane of the post-acrosome. The role of the Catsper channel in boar spermatozoa was investigated by extending the spermatozoa in media containing different calcium (Ca(2+)) availability and exposure to the capacitation-trigger bicarbonate, to progesterone or CatSper inhibitors (Mibefradil and NNC 55-0396), separately or sequentially, at physiological and toxicological doses. Extracellular Ca(2+) availability, combined with bicarbonate exposure (capacitation-inducing conditions) decreased sperm motility, similarly to when spermatozoa incubated in capacitation-inducing conditions was exposed to Mibefradil and NNC 55-0396. Exposure of these spermatozoa to progesterone did not cause significant changes in sperm motility and nor did it revert its decrease induced by CatSper antagonists. In conclusion, the CatSper channel regulates sperm motility during porcine capacitation-related events in vitro.

    Ort, förlag, år, upplaga, sidor
    Elsevier, 2017
    Nyckelord
    CatSper, Spermatozoa, Capacitation, Sperm motility, Boar
    Nationell ämneskategori
    Biologiska vetenskaper
    Identifikatorer
    urn:nbn:se:liu:diva-134398 (URN)10.1016/j.repbio.2017.01.001 (DOI)000397370900009 ()28077244 (PubMedID)2-s2.0-85009212156 (Scopus ID)
    Anmärkning

    Funding agencies: The Swedish Research council VR, Stockholm [521-2011-6553]; Research Council FORMAS, Stockholm [221-2011-512]; FORSS (Forskningsradet i Sydostra Sverige, Sweden [473121]

    Tillgänglig från: 2017-02-23 Skapad: 2017-02-23 Senast uppdaterad: 2017-05-29Bibliografiskt granskad
    3. The mu (µ) and delta (δ) opioid receptors modulate boar sperm motility
    Öppna denna publikation i ny flik eller fönster >>The mu (µ) and delta (δ) opioid receptors modulate boar sperm motility
    2016 (Engelska)Ingår i: Molecular Reproduction and Development, ISSN 1040-452X, E-ISSN 1098-2795, Vol. 83, nr 8, s. 724-734Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Endogenous and exogenous opioids modulate reproductive functions in target cells via opioid receptors (µ, δ, and κ). Sperm motility is a metric of gamete functionality, and serves as a suitable parameter for in vitro drug-induced toxicity assays. This study identifies the presence and location of opioid receptors in pig spermatozoa as well as their functional response after in vitro challenge with known agonists (morphine [µ]; [D-Pen 2,5]-enkephanile [δ]; and U 50488 [κ]) and antagonists (naloxone [µ]; naltrindole [δ]; and nor-binaltrorphimine [κ]). Only the µ- and δ-opioid receptors were present in the sperm plasma membrane, overlying the acrosome, neck, and principal piece. Challenge experiments with agonists and antagonists identified both µ- and δ-opioid receptors as regulators of sperm kinematics, wherein µ maintains or increases sperm movement whereas δ decreases sperm motility over time. This article is protected by copyright. All rights reserved.

    Ort, förlag, år, upplaga, sidor
    John Wiley & Sons, 2016
    Nyckelord
    opioids, membrane receptors, kinematics, pig
    Nationell ämneskategori
    Utvecklingsbiologi
    Identifikatorer
    urn:nbn:se:liu:diva-130181 (URN)10.1002/mrd.22675 (DOI)000387014800007 ()27391529 (PubMedID)
    Anmärkning

    Funding agencies: Swedish Research Council VR, Stockholm [521-2011-6553]; Research Council FORMAS, Sweden [221-2011-512]; Research Council in Southeast Sweden (FORSS), Sweden [378091/31297]

    Tillgänglig från: 2016-07-14 Skapad: 2016-07-14 Senast uppdaterad: 2017-11-28Bibliografiskt granskad
    4. Boar spermatozoa successfully predict mitochondrial modes of toxicity: Implications for drug toxicity testing and the 3R principles
    Öppna denna publikation i ny flik eller fönster >>Boar spermatozoa successfully predict mitochondrial modes of toxicity: Implications for drug toxicity testing and the 3R principles
    Visa övriga...
    2015 (Engelska)Ingår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 29, nr 3, s. 582-591Artikel i tidskrift (Refereegranskat) Published
    Abstract [en]

    Replacement of animal testing by in vitro methods (3-R principles) requires validation of suitable cell models, preferably obtained non-invasively, defying traditional use of explants. Ejaculated spermatozoa are highly dependent on mitochondrial production and consumption of ATP for their metabolism, including motility display, thus becoming a suitable model for capturing multiple modes of action of drugs and other chemicals acting via mitochondrial disturbance. In this study, a hypothesis was tested that the boar spermatozoon is a suitable cell type for toxicity assessment, providing a protocol for 3R-replacement of animals for research and drug-testing. Boar sperm kinetics was challenged with a wide variety of known frank mito-toxic chemicals with previously shown mitochondrial effects, using a semi-automated motility analyser allied with real-time fluorescent probing of mitochondrial potential (MitoTracker and JC-1). Output of this sperm assay (obtained after 30 min) was compared to cell viability (ATP-content, data obtained after 24-48 h) of a hepatome-cell line (HepG2). Results of compound effects significantly correlated (P less than 0.01) for all sperm variables and for most variables in (HepG2). Dose-dependent decreases of relative ATP content in HepG2 cells correlated to sperm speed (r= 0.559) and proportions of motile (r = 0.55) or progressively motile (r = 0.53) spermatozoa. The significance of the study relies on the objectivity of computerized testing of sperm motility inhibition which is comparable albeit of faster output than somatic cell culture models. Sperm suspensions, easily and painlessly obtained from breeding boars, are confirmed as suitable biosensors for preclinical toxicology screening and ranking of lead compounds in the drug development processes.

    Ort, förlag, år, upplaga, sidor
    Elsevier, 2015
    Nyckelord
    Sperm; Motility; Mitochondria; Drug; Toxicity; Boar
    Nationell ämneskategori
    Klinisk medicin
    Identifikatorer
    urn:nbn:se:liu:diva-117655 (URN)10.1016/j.tiv.2015.01.004 (DOI)000352050100019 ()25624015 (PubMedID)
    Anmärkning

    Funding Agencies|Swedish Research Council (VR); Research Council Formas, Stockholm, Sweden

    Tillgänglig från: 2015-05-12 Skapad: 2015-05-06 Senast uppdaterad: 2017-12-04
  • 13.
    Vicente Carrillo, Alejandro
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Alvarez-Rodriguez, Manuel
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    The mu (µ) and delta (δ) opioid receptors modulate boar sperm motility2016Ingår i: Molecular Reproduction and Development, ISSN 1040-452X, E-ISSN 1098-2795, Vol. 83, nr 8, s. 724-734Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Endogenous and exogenous opioids modulate reproductive functions in target cells via opioid receptors (µ, δ, and κ). Sperm motility is a metric of gamete functionality, and serves as a suitable parameter for in vitro drug-induced toxicity assays. This study identifies the presence and location of opioid receptors in pig spermatozoa as well as their functional response after in vitro challenge with known agonists (morphine [µ]; [D-Pen 2,5]-enkephanile [δ]; and U 50488 [κ]) and antagonists (naloxone [µ]; naltrindole [δ]; and nor-binaltrorphimine [κ]). Only the µ- and δ-opioid receptors were present in the sperm plasma membrane, overlying the acrosome, neck, and principal piece. Challenge experiments with agonists and antagonists identified both µ- and δ-opioid receptors as regulators of sperm kinematics, wherein µ maintains or increases sperm movement whereas δ decreases sperm motility over time. This article is protected by copyright. All rights reserved.

  • 14.
    Vicente Carrillo, Alejandro
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Hälsouniversitetet.
    Edebert, I.
    Karlbergsvägen 83 B, Stockholm, Sweden.
    Garside, H.
    AstraZeneca Research and Dev, England.
    Cotgreave, I.
    Karolinska Institute, Sweden; Karolinska Institute, Sweden.
    Rigler, R.
    Karolinska Institute, Sweden.
    Loitto, Vesa
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Hälsouniversitetet.
    Magnusson, Karl-Eric
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för mikrobiologi och molekylär medicin. Linköpings universitet, Hälsouniversitetet.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Boar spermatozoa successfully predict mitochondrial modes of toxicity: Implications for drug toxicity testing and the 3R principles2015Ingår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 29, nr 3, s. 582-591Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Replacement of animal testing by in vitro methods (3-R principles) requires validation of suitable cell models, preferably obtained non-invasively, defying traditional use of explants. Ejaculated spermatozoa are highly dependent on mitochondrial production and consumption of ATP for their metabolism, including motility display, thus becoming a suitable model for capturing multiple modes of action of drugs and other chemicals acting via mitochondrial disturbance. In this study, a hypothesis was tested that the boar spermatozoon is a suitable cell type for toxicity assessment, providing a protocol for 3R-replacement of animals for research and drug-testing. Boar sperm kinetics was challenged with a wide variety of known frank mito-toxic chemicals with previously shown mitochondrial effects, using a semi-automated motility analyser allied with real-time fluorescent probing of mitochondrial potential (MitoTracker and JC-1). Output of this sperm assay (obtained after 30 min) was compared to cell viability (ATP-content, data obtained after 24-48 h) of a hepatome-cell line (HepG2). Results of compound effects significantly correlated (P less than 0.01) for all sperm variables and for most variables in (HepG2). Dose-dependent decreases of relative ATP content in HepG2 cells correlated to sperm speed (r= 0.559) and proportions of motile (r = 0.55) or progressively motile (r = 0.53) spermatozoa. The significance of the study relies on the objectivity of computerized testing of sperm motility inhibition which is comparable albeit of faster output than somatic cell culture models. Sperm suspensions, easily and painlessly obtained from breeding boars, are confirmed as suitable biosensors for preclinical toxicology screening and ranking of lead compounds in the drug development processes.

  • 15.
    Vicente-Carrillo, Alejandro
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Ekwall, H
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Álvarez-Rodríguez, Manuel
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Membrane Stress During Thawing Elicits Redistribution of Aquaporin 7 But Not of Aquaporin 9 in Boar Spermatozoa2016Ingår i: Reproduction in domestic animals, ISSN 0936-6768, E-ISSN 1439-0531, Vol. 51, nr 5, s. 665-679Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Freezing of boar spermatozoa includes the cryoprotectant glycerol, but renders low cryosurvival, owing to major changes in osmolarity during freezing/thawing. We hypothesize that aquaporins (AQPs) 7 and 9 adapt their membrane domain location to these osmotic challenges, thus maintaining sperm homeostasis. Western blotting (WB) and immunocytochemistry (ICC) at light and electron microscope levels with several commercial primary antibodies and protocols explored AQP location on cauda epididymal and ejaculated spermatozoa (from different fractions of the ejaculate), unprocessed, extended, chilled and frozen-thawed. Although differences in WB and ICC labelling were seen among antibodies, AQP-7 was conspicuously located in the entire tail and cytoplasmic droplet in caudal spermatozoa, being restricted to the mid-piece and principal piece domains in ejaculated spermatozoa. AQP-9 was mainly localized in the sperm head in both caudal and ejaculated spermatozoa. While unaffected by chilling (+5°C), freezing and thawing of ejaculated spermatozoa clearly relocated the head labelling of AQP-7, but not that of AQP-9. In vitro mimicking of cell membrane expansion during quick thawing maintained the localization of AQP-9 but relocated AQP-7 towards the acrosome. AQP-7, but not AQP-9, appears as a relevant marker for non-empirical studies of sperm handling.

  • 16.
    Vicente-Carrillo, Alejandro
    et al.
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för kliniska vetenskaper. Linköpings universitet, Medicinska fakulteten.
    Álvarez-Rodríguez, Manuel
    Linköpings universitet, Medicinska fakulteten. Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa.
    Rodríguez-Martínez, Heriberto
    Linköpings universitet, Medicinska fakulteten. Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för barns och kvinnors hälsa.
    The CatSper channel modulates boar sperm motility during capacitation.2017Ingår i: Reproductive biology, ISSN 2300-732X, Vol. 17, nr 1, s. 69-78Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The cation channel of sperm (CatSper) comprises four transmembrane subunits specifically expressed in human, equine, murine and ovine spermatozoa, apparently implicated in capacitation, hyperactivation and acrosome exocytosis. Western blotting and immunocytochemistry showed hereby that CatSper subunits are also present in boar spermatozoa, primarily over the sperm neck, tail and cytoplasmic droplets; albeit CatSper -1 presented in addition some distribution over the membrane of the acrosome and CatSper -2 and -4 over the membrane of the post-acrosome. The role of the Catsper channel in boar spermatozoa was investigated by extending the spermatozoa in media containing different calcium (Ca(2+)) availability and exposure to the capacitation-trigger bicarbonate, to progesterone or CatSper inhibitors (Mibefradil and NNC 55-0396), separately or sequentially, at physiological and toxicological doses. Extracellular Ca(2+) availability, combined with bicarbonate exposure (capacitation-inducing conditions) decreased sperm motility, similarly to when spermatozoa incubated in capacitation-inducing conditions was exposed to Mibefradil and NNC 55-0396. Exposure of these spermatozoa to progesterone did not cause significant changes in sperm motility and nor did it revert its decrease induced by CatSper antagonists. In conclusion, the CatSper channel regulates sperm motility during porcine capacitation-related events in vitro.

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