Epithelial cadherin is present in bovine oviduct epithelial cells and gametes,and is involved in fertilization-related events

Fertilization is a calcium-dependent process that involves sequential cell–cell adhesion events of spermatozoa with oviduct epithelial cells (OECs) and with cumulus-oocyte complexes (COCs). Epithelial cadherin (E-cadherin) participates in calcium-dependent somatic cell adhesion; the adaptor protein β-catenin binds to the E-cadherin cytoplasmic domain and links the adhesion protein to the cytoskeleton. The study was conducted to immunodetect E-cadherin and β-catenin in bovine gametes and oviduct (tissue sections and OEC monolayers), and to assess E-cadherin participation in fertilization-related events. Epithelial cadherin was found in spermatozoa, oocytes, cumulus cells, and OEC. In acrosome-intact noncapacitated spermatozoa, E-cadherin was mainly localized in the apical ridge and acrosomal cap (E1-pattern; 84 ± 9%; mean ± standard deviation of the mean). After sperm treatment with heparin to promote capacitation, the percentage of cells with E1-pattern (56 ± 12%) significantly decreased; concomitantly, the percentage of spermatozoa depicting an E-cadherin staining pattern similar to E1-pattern but showing a signal loss in the acrosomal cap (E2-pattern: 40 ± 11%) increased. After l-α-lysophosphatidylcholine–induced acrosome reaction, E-cadherin signal was mainly localized in the inner acrosomal membrane (E3-pattern: 67 ± 22%). In IVM COC, E-cadherin was immunodetected in the plasma membrane of cumulus cells and oocytes, but was absent in the polar body. The 120 KDa mature protein form was found in protein extracts from spermatozoa, oocytes, cumulus cells, and OEC. β-Catenin distribution followed E-cadherin's in all cells evaluated. Epithelial cadherin participation in cell–cell interaction was evaluated using specific blocking monoclonal antibody DECMA-1. Sperm incubation with DECMA-1 impaired sperm–OEC binding (the number of sperm bound to OEC: DECMA-1 = 6.7 ± 6.1 vs. control = 29.6 ± 20.1; P < 0.001), fertilization with COC (% fertilized COC: DECMA-1 = 68.8 ± 10.4 vs. control = 90.7 ± 3.1; P < 0.05) or denuded oocytes (% fertilized oocytes: DECMA-1 = 57.0 ± 15.2 vs. control = 89.2 ± 9.8; P < 0.05) and binding to the oolemma (the number of sperm bound to oolemma: DECMA-1 = 2.2 ± 1.1 vs. control = 11.1 ± 4.8; P < 0.05). This study describes, for the first time, the presence of E-cadherin in bovine spermatozoa, COC, and OEC, and shows evidence of its participation in sperm interaction with the oviduct and the oocyte during fertilization.     

Historical perspectives and recent research on superovulation in cattle

Superovulation protocols have evolved greatly over the past 40 to 50 years. The development of commercial pituitary extracts and prostaglandins in the 1970s, and partially purified pituitary extracts and progesterone-releasing devices in the 1980s and 1990s have provided for the development of many of the protocols that we use today. Furthermore, the knowledge of follicular wave dynamics through the use of real-time ultrasonography and the development of the means by which follicular wave emergence can be controlled have provided new practical approaches. Although some embryo transfer practitioners still initiate superstimulatory treatments during mid-cycle in donor cows, the elective control of follicular wave emergence and ovulation has had a great effect on the application of on-farm embryo transfer, especially when large groups of donors need to be superstimulated at the same time. The most common treatment for the synchronization of follicular wave emergence for many years has been estradiol and progestins. In countries where estradiol cannot be used, practitioners have turned to alternative treatments for the synchronization of follicle wave emergence, such as mechanical follicle ablation or the administration of GnRH to induce ovulation. An approach that has shown promise is to initiate FSH treatments at the time of the emergence of the new follicular wave after GnRH-induced ovulation of an induced persistent follicle. Alternatively, it has been suggested recently that it might be possible to ignore follicular wave status, and by extending the treatment protocol, induce small antral follicles to grow and superovulate. Recently, the mixing of FSH with sustained release polymers or the development of long-acting recombinant FSH products have permitted superstimulation with a single or alternatively, two gonadotropin treatments 48 hours apart, reducing the need for animal handling during superstimulation. Although the number of transferable embryos per donor cow superstimulated has not increased, the protocols that are used today have increased the numbers of transferable embryos recovered per unit time and have facilitated the application of on-farm embryo transfer programs. They are practical, easy to administer by farm personnel, and more importantly, they eliminate the need for detecting estrus.     

Electron microscopy morphology of the mitochondrial network in gliomas and their vascular microenvironment

Gliomas still represent a serious and discouraging brain tumor; despite of the diversity of therapeutic modalities, the prognosis for patients is still poor. Understanding the structural and functional characteristics of the vascular microenvironment in gliomas is essential for the design of future therapeutic strategies. This review describes and analyzes the electron microscopy morphology of the mitochondrial network in human gliomas and their vascular microenvironment. Heterogenous mitochondrial network alterations in glioma cells and in microvascular environment are implicated directly and indirectly in the processes linked to hypoxia-tolerant and hypoxia-sensitive cells phenotype, effects of the hypoxia-inducible factor-1α, increased expression of several glycolytic protein isoforms as well as fatty acid synthase, and survivin. The prevalent existence of partial and total cristolysis observed suggests that oxidative phosphorylation is severely compromised. A mixed therapy emerged as the most appropriate.     

The intracellular distal tail of the Na+/H+ exchanger NHE1 is intrinsically disordered: implications for NHE1 trafficking

Intrinsic disorder is important for protein regulation, yet its role in regulation of ion transport proteins is essentially uninvestigated. The ubiquitous plasma membrane carrier protein Na(+)/H(+) Exchanger isoform 1 (NHE1) plays pivotal roles in cellular pH and volume homeostasis, and its dysfunction is implicated in several clinically important diseases. This study shows, for the first time for any carrier protein, that the distal part of the C-terminal intracellular tail (the cdt, residues V686-Q815) from human (h) NHE1 is intrinsically disordered. Further, we experimentally demonstrated the presence of a similar region of intrinsic disorder (ID) in NHE1 from the teleost fish Pleuronectes americanus (paNHE1), and bioinformatic analysis suggested ID to be conserved in the NHE1 family. The sequential variation in structure propensity as determined by NMR, but not the amplitude, was largely conserved between the h- and paNHE1cdt. This suggests that both proteins contain molecular recognition features (MoRFs), i.e., local, transiently formed structures within an ID region. The functional relevance of the most conserved MoRF was investigated by introducing a point mutation that significantly disrupted the putative binding feature. When this mutant NHE1 was expressed in full length NHE1 in AP1 cells, it exhibited impaired trafficking to the plasma membrane. This study demonstrated that the distal regulatory domain of NHE1 is intrinsically disordered yet contains conserved regions of transient structure. We suggest that normal NHE1 function depends on a protein recognition element within the ID region that may be linked to NHE1 trafficking via an acidic ER export motif.     

Solid character of membrane ceramides: a surface rheology study of their mixtures with sphingomyelin

The compression and shear viscoelasticities of egg-ceramide and its mixtures with sphingomyelin were investigated using oscillatory surface rheology performed on Langmuir monolayers. We found high values for the compression and shear moduli for ceramide, compatible with a solid-state membrane, and extremely high surface viscosities when compared to typical fluid lipids. A fluidlike rheological behavior was found for sphingomyelin. Lateral mobilities, measured from particle tracking experiments, were correlated with the monolayer viscosities through the usual hydrodynamic relationships. In conclusion, ceramide increases the solid character of sphingomyelin-based membranes and decreases their fluidity, thus drastically decreasing the lateral mobilities of embedded objects. This mechanical behavior may involve important physiological consequences in biological membranes containing ceramides.     

Autofluorescence of the fungus <Emphasis Type="Italic">Morchella conica</Emphasis> var. <Emphasis Type="Italic">rigida</Emphasis>

Autofluorescence (primary fluorescence (AF)) of fruiting bodies and stems of the fungus Morchella conica var. rigida was studied by fluorescence microscopy including sporangia and ascospores. The ascospores were characterized by a weak green–yellow AF at blue excitation. Using a green excitation, no AF was observed. The hyphae located under the layer of asci with ascospores exhibited a higher primary fluorescence, namely their walls that had green-yellow color at blue excitation. Also, their red AF observed when a green excitation was used was significant. Similarly, the hyphae located in the fungal stem exhibited a significant AF, especially their walls when the blue light was used for excitation. In addition, large, yellow-to-yellow/green, oval-to-round bodies with strong fluorescence were detected whose morphological equivalents were not clearly visible in the white halogen light. The AF of the fungus M. conica var. rigida was lower compared with the other higher fungi studied so far.