Notch signaling regulates nuclear androgen receptor AR and membrane androgen receptor ZIP9 in mouse Sertoli cells.

BACKGROUND: Notch signaling pathway is involved in contact-dependent communication between the cells of seminiferous epithelium, and its proper activity is important for undisturbed spermatogenesis. OBJECTIVES: The aim was to assess the effect of Notch pathway inhibition on the expression of nuclear (AR) and membrane (ZIP9) androgen receptors and androgen-regulated genes, claudin-5 and claudin-11, in TM4 mouse Sertoli cell line. MATERIALS AND METHODS: DAPT (γ-secretase inhibitor) treatment and recombination signal binding protein silencing were employed to reduce Notch signaling, whereas immobilized ligands were used to activate Notch pathway in TM4 cells. To reveal specific effect of each androgen receptor, AR or ZIP9 silencing was performed. RESULTS: Notch pathway inhibition increased the expression of AR and ZIP9 mRNA and proteins (p < 0.01; p < 0.05) in TM4 cells, whereas incubation with Notch ligands, rDLL1 or rJAG1, reduced AR (p < 0.01; p < 0.001) and ZIP9 (p < 0.05; p < 0.01) expressions, respectively. Testosterone enhanced the expression of both receptors (p < 0.05; p < 0.01). Androgen-regulated claudin-5 and claudin-11 (p < 0.01; p < 0.001) and cAMP (p < 0.001) were elevated in Notch-inhibited cells, while activation of Notch signaling by DLL1 or JAG1 reduced claudin-11 or claudin-5 level (p < 0.01; p < 0.001), respectively. DISCUSSION: Our findings indicate opposite effect of Notch and androgen signaling on the expression of androgen receptors in TM4 cells. We demonstrated that AR expression is regulated by DLL1-mediated Notch signaling, whereas JAG1 is involved in the regulation of ZIP9. The expression of both claudins and cAMP production is under inhibitory influence of Notch pathway. The effects of Notch signaling on claudin-5 and claudin-11 expression are mediated by ZIP9 and AR, respectively. CONCLUSION: Notch signaling may be considered as an important pathway controlling Sertoli cell physiology, and its alterations may contribute to disturbed response of Sertoli cells to androgens.

Markers of the progression of complications in patients with type 2 diabetes: a one-year longitudinal study.

Hyperglycemia induces tissue damage and complications by mechanisms that produce advanced glycation end-products (AGEs) and inflammation.To investigate the factors associated with the progression of complications in Type 2 diabetes patients.We recruited 157 patients (110 women and 47 men) with diabetes for more than 5 years who were non-smokers and did not have current infections or chronic diseases. Patients were grouped according to neuropathy, nephropathy, and retinopathy status: without (I), slight or moderate (II), and severe complications (III). We measured glucose, lipids and HbA1c, low molecular weight AGEs (LMW AGEs), high sensitivity C-reactive protein (CRP), TNF-α, IL-6, and malondialdehyde (MDA). Patients were re-evaluated 1 year later.Patients were 52.2±6.8 years old with 11.0±4.9 years since diagnosis. After 1 year, circulating AGEs increased (p<0.0001) and eGFR decreased (p<0.0007) in groups II and III. IL-6 and MDA decreased in groups I and II. CRP (p<0.029) and AGEs (p<0.0001) increased in group II. At baseline in group I, TNF-α levels were higher (p<0.002) in patients who later developed complications. In group II, TNF-α levels (p<0.015) and microalbuminuria (p<0.00004) were higher in patients whose complications progressed. Logistic regression analysis showed that complication progress was significantly associated with log(albuminuria) (p<0.004) and log(TNF-α) (p<0.008). In the total group, AGEs were associated with age (p<0.024) and HbA1c (p<0.026).Our results suggest that baseline TNF-α is an important predictor of complication progression in Type 2 diabetes patients. AGEs also increased during the deterioration of renal function after 1 year of follow-up observation.

Role of the mesonephros as a transient haematopoietic organ in the bovine embryo.

During vertebrate embryogenesis, haematopoietic stem cells (HSC) arise in the aorta-gonads-mesonephros (AGM) region. In the present study, we examined serial sections of 22-34 days post-insemination (dpi) bovine embryos, a species with a well-developed and functional mesonephros. We describe the temporo-spatial distribution of presumptive c-kit positive HSC and the occurrence of haematopoietic foci in the mesonephros and fetal liver using specific antibodies directed against haematopoietic cell markers and conventional electron microscopy. In the mesonephros, presumptive HSC were found at 23-24 dpi in the blood stream, in the endothelial lining of the filtering capillaries and in the septal stroma of the cranial part of the mesonephros, the mesonephric giant corpuscle (MGC), suggesting a colonalization via the blood stream from the haematopoietic clusters of the dorsal aorta. From 25 to 30 dpi, presumptive HSC predominate in the septal stroma of the MGC, were they first expand but then decline and disappear following 32 dpi. In parallel, we found ongoing erythropoiesis and myelopoiesis starting in the MGC at 24 dpi and extending during the complete observation period. In the embryonic liver, colonization with presumptive c-kit positive HSC occurs slightly later, at 25 dpi. Active formation of blood cells in the liver increases following 30 dpi. In conclusion, the mesonephros of bovine embryos, in particular its MGC, functions as a primitive haematopoietic organ, temporarily intercalated between extraembryonic erythropoiesis and haematopoiesis within in the fetal liver, thus recapitulating for a short period a phylogenetically old site of blood formation.

Exposure to organic solvents and cytogenetic damage in exfoliated cells of the buccal mucosa from shoe workers.

PURPOSE: We determined the solvents mainly used in shoe making and their genotoxic effects. METHODS: Thirty-four exposed shoe workers and 34 unexposed control subjects, paired by age and sex, were compared. Occupational exposure was determined by using monitors 3M. Solvents were assessed by gas chromatography. Exfoliated buccal cells were obtained from each subject to determine the incidence of micronuclei and other nuclear abnormalities. One thousand cells were counted in each subject. RESULTS: Solvents detected were acetone, ethyl acetate, methyl ethyl ketone, and toluene. The incidence of nuclear abnormalities was significatively higher in the exposed group when compared to the control group. A positive relationship between the incidence of micronuclei and the toluene concentration in the environment was found. CONCLUSIONS: Toluene shows an important genotoxic effect. As the micronuclei test is an effective, fast, simple and low cost biomarker to identify cytogenetic effects, we suggest its utilization as a preventive test of genotoxicity.

Desarrollo de pan integral con soya, chía, linaza y ácido fólico como alimento funcional para la mujer / Integral bread development with soybean, chia, linseed, and folic acid as a functional food for women

Six bread formulations were developed, using different proportions of whole-wheat flour, chia seeds and flaxseed flour. All of our formulations were added with folic acid. Sensorial and texture evaluations were performed, showing good acceptance of the products. Proximal chemical analysis was carried out; in addition, the following parameters were determined: calcium, phosphorus, total dietary fiber, folic acid, water hydration capacity, Glucose Dialysis Retardation Index (GDRI) and fatty acids. The results obtained showed higher protein levels in the developed breads (23.23-30.24 (g/100g dry matter) as compared to a control (21.00% of proteins in bread elaborated without chia or flaxseed). Furthermore, the breads contained 10.07-12.15 of lipids (g/100g dry matter) (linoleic acid: 2.43-4.05%; linolenic acid: 1.12-4.46 %; oleic acid: 2.93-6.13 %), GDRI values were between 89.1 and 98.1 % and folic acid was in the range 699.44 - 991.3 (microg/100g dry matter). The same parameters were determined in the chia seed and in the flaxseed flour. It was concluded that; due to their high levels of protein, insaturated fatty acids (omega-3 and omega-6), dietary fiber and folic acid, these breads have a high nutritional value, so they could have special benefits for woman.

Immunophenotyping and spatio-temporal distribution of aortic cell clusters in the bovine embryo.

In the present study the temporal and spatial appearance of aortic cell clusters in bovine embryos is described. Aorta-associated c-kit-positive cell clusters can be observed first in 23 days post inseminationem (dpi) bovine embryos and disappear after 34 dpi. For the first time, it was shown that the immunophenotype of these aortic cluster cells changes during embryonic development. Aortic cell clusters are c-kit+/CD45-/STA-, when they are first detected in the 23 dpi embryo, and acquire a c-kit+/CD45+/STA- phenotype in 27-29 embryos and a c-kit+/CD45+/STA+ immunophenotype in 32-34-day-old specimens. Cell clusters are most prominent in the vicinity of lateral and ventral aortic branches, but rare in omphalomesenteric arteries and absent in Aa. umbilicales. Free c-kit-positive cells in an intravasal position are common, suggesting separation from the clusters in order to colonize subsequent hematopoietic organs, i.e., the liver and the mesonephros. Transmission electron microscopic analysis reveals the existence of primitive desmosomes between the clusters cells and adjacent endothelial cells as well as a fine basal lamina as a demarcation between the cluster cells and underlying mesenchymal cells. Material resembling extracellular matrix is found in large vacuoles in cluster cells of 23 dpi embryos. Immunocytochemistry reveals an intense accumulation of heparan sulfate proteoglycan and collagen IV in the aortic wall at the sites where cell clusters are attached. These observations suggest that the hematopoietic cell clusters induce the formation of a specific microenvironment within the aortic wall.

Advanced glycosylation end products in skin, serum, saliva and urine and its association with complications of patients with type 2 diabetes mellitus.

BACKGROUND: The accumulation of advanced glycation end products (AGEs) has a key role in the pathophysiology of diabetes complications. Comparison of AGEs measurement in serum, skin, saliva and urine has not been reported. AIMS: To compare AGEs in serum, skin, saliva and urine in patients with Type 2 diabetes mellitus, with complications at different stages. MATERIALS AND METHODS: We examined 50 patients with Type 2 diabetes mellitus (40 women and 10 men) grouped according to the progression of neuropathy, nephropathy and retinopathy. The AGEs content in serum, skin, saliva and urine was measured by spectrofluorometry HPLC. RESULTS: The patients had a mean age of 56.5 +/- 7.7 yr and 12.8 +/- 6.7 yr since diagnosis. AGEs in skin correlated with years since diagnosis (p = 0.0005). AGEs in serum, skin and saliva increased with the progression of complications, nevertheless, in urine a trend to diminution was found. In the group with end-stage renal disease (ESRD), AGEs in serum increased in greater proportion. In order to account for the decreased AGEs clearance, we corrected the values for creatinine levels, and AGEs in skin gave a better association with complications. CONCLUSIONS: The AGEs measurement in skin, serum and saliva are useful to evaluate diabetes complications. AGEs in skin are associated with years since diagnosis of diabetes. Correction for renal function might discriminate AGEs in situ formation from accumulation.

Morphology and innervation pattern of the feline urogenital junction.

The feline urogenital junction is situated between the extratesticular rete and the spacious initial segments of the efferent ductules. The rete epithelium is cuboidal to low columnar. The rete cells forming the junction rest on a wavy basal lamina, display deep mutual invaginations, possess central nuclei with several infoldings and form a distinct border with the columnar epithelial cells of the initial segments of the ductuli efferentes. The epithelium of the initial segments is composed of ciliated cells and non-ciliated principal cells. The latter are the dominating type and characterized by an apical brush-border and a supranuclear endocytotic apparatus. The stroma of the extratesticular rete contains an abundance of collagen whereas contractile cells are here generally absent. In contrast, the initial segments of the efferent ductules are surrounded by elastic fibres and a layer of contractile cells. All nerves for the feline urogenital junction come from the nervus spermaticus superior. In the epididymal head, small nerve bundles deviate into the septa between the ductules. Single fibres establish a dense network within the muscular coat of the ductuli. At the transition to the extratesticular rete, this network ends abruptly. Nerve fibres in the confines of the rete are associated with blood vessels or proceed to the testicular interior, but establish no relationships with the rete epithelium or the myofibroblasts of the mediastinum. The nervous network in the walls of the efferent ductules and their initial segments is not only composed of sympathetic but also parasympathetic, non-myelinated fibres. Particularly noteworthy is the abundance of calcitonin gene-related peptide (CGRP)- and substance P (SP)-containing axons around the initial segments. Both neuroproteins are consistent markers for sensory neurones. Taken together, it can be assumed that the entry of seminal fluid and spermatozoa into the efferent ductules is controlled by a regulatory nervous chain provided with afferent and efferent components.

Establishment of the urogenital junction in the male bovine embryo: an ultrastructural study.

The ultrastructure of the developing extratesticular rete testis, the efferent ductules and the establishment of the urogenital junction were studied in bovine embryos and fetuses of 41 through 95 days post conceptionem. The efferent ductules originate as a new set of secondary mesonephric tubules from the dorsal aspect of the nephric giant corpuscle and grow in the direction of the Wolffian duct. Cytological differentiation of the efferent ductules proceeds in a proximo--distal direction. At about 50-60 days, the simple columnar epithelium of the proximal portions of the efferent ductules already consists of the two typical cell types, i.e. reabsorptive principal cells with an endocytotic apparatus and a brush-border and ciliated cells. The lumen of the proximal portion is temporarily filled with intraductular blood vessels and perivascular tissue which may represent vestigial rudiments of glomeruli associated with the efferent ductules. At 50 to 60 days, the extratesticular rete still has a blastema--like appearance and consists of irregular cells with abundant glycogen. Extensions of the extratesticular rete come into contact with the efferent ductules and create the first end-to-side anastomoses with the latter. Somewhat later, the separating basal laminas vanish and invading rete cells intermingle with the epithelium of the efferent ductules, thus establishing the urogenital junction.

The economics of health insurance plan design.

In developing insurance products, health insurers have often failed to incorporate member-level economic decision making into the underlying plan design. Instead, health insurers have either completely isolated members from economic choices within a restricted network or simply shifted costs without providing members with an opportunity to more efficiently purchase health care services. In either approach, the plan designs do not provide members with a rational economic framework to make efficient health care purchases. In order to analyze the economic framework in traditional plan designs, this article first outlines the characteristics of an efficient economic transaction and then compares these characteristics with traditional insurance plans. The article concludes by suggesting new plan designs that achieve more economically efficient outcomes through the application of episode of care technology.

The nerve distribution in the testis of the cat.

The autonomous innervation of the feline testis was investigated by immunohistochemistry and a modified acetylcholinesterase technique. The nerves reach the testis mainly by two routes: (1) with testicular artery and pampiniform plexus to the cranial extremity (funicular contribution), (2) from the epididymal tail to the caudal extremity (caudal contribution). Within the tunica albuginea the funicular contribution supplies the cranial two thirds, whereas the caudal third of the tunica receives its nerves via the ligamentous connection between testis and epididymal tail. The nerve bundles accompanying the testicular artery give branches to the arterial wall and the pampiniform plexus. When reaching the cranial testicular pole the bundles separate; the majority of them pass into the centrally located mediastinum testis, another large portion enters the tunica albuginea, particularly on its epididymal side. The septula testis are innervated from both sides, that is from the mediastinum and from the tunica albuginea. In the cat, contrary to other mammals, all septula are innervated. Furthermore, nerve fibers occur regularly within the testicular lobules. Generally, the testicular nerves of the cat are unmyelinated and mainly vascular nerves, but fibers are also found within the connective tissue compartments of the testis. The vast majority of all autonomous testicular nerves are postjunctional sympathetic fibers. Terminal ramifications of cholinergic fibers are exclusively observed in the wall of medium-sized arterioles within mediastinum, septula and lobuli testis. Neuropeptide Y is the most frequent peptidergic transmitter in feline testicular vascular plexuses. The amount of calcitonin gene-related peptide-positive fibers is also remarkably high in the testis, but prefers a location within the stroma of the tunica albuginea, mediastinum and septula. In the cat, Leydig cells occur not only in intertubular locations, but also as intratunical and mediastinal Leydig cells. In all three localizations solitary nerve fibers are observed between Leydig cell groups. These fibers are generally dopamin-beta-hydroxylase- and tyrosine hydroxylase-positive, some contain calcitonin gene-related peptide and, very few, substance P.

Morphogenesis of the bovine rete testis: extratesticular rete, mesonephros and establishment of the definitive urogenital junction.

The development of the extratesticular rete, the regression of the mesonephros and the establishment of the urogenital junction between rete testis and efferent ductules were investigated in 67 bovine embryos and fetuses collected in the period from day 29 through day 250 post conception. The results were obtained by immunohistochemistry and by the study of semithin sections. At about day 30, the large mesonephros contains a peculiar Malpighian body in its cranial part, generally referred to as the mesonephric giant corpuscle, which is connected to the Wolffian duct by a series of well-developed and functioning mesonephric tubules. This set of primary mesonephric tubules, however, will not participate in the formation of the definitive urogenital junction, but will regress and soon disappear completely. The efferent ductules in the bovine are represented by another set of secondary mesonephric tubules that grow out from the dorsal aspect of the mesonephric giant corpuscle at about day 50. Transiently, the lumina of the sprouting efferent ductules are plugged by invading intraductular blood vessels, probably representing rudimentary glomeruli. The proximal portions of the newly-formed efferent ductules establish side-to-end contacts with extensions of the extratesticular rete that has bypassed the regressing giant corpuscle. At 85 days, the efferent ductules have reached the Wolffian duct and open into it. At 150 days, the channels of the extratesticular rete display a patent lumen and now form end-to-end anastomoses with the efferent ductules. The proliferating mesenchymal cells surrounding the epithelia of the efferent ductules have arranged in several concentric layers at about 85 days. These mesenchymal cells are the precursors of the periductular musculature and are reached by the first nerve fibers at about day 130.

Morphogenesis of the bovine rete testis: the intratesticular rete and its connection to the seminiferous tubules.

The development of the intragonadal rete testis and the establishment of the connection between seminiferous and straight testicular tubules was studied using ultrastructural and histochemical methods in 60 bovine embryos and fetuses ranging from day 39 through day 225 post conceptionem. The methodology included a modified acetylcholinesterase (AChE) reaction as a selective marker for pre-Sertoli cells and a modified microsomal aminopeptidase (MAP) reaction as a selective marker for the epithelia of rete testis and straight testicular tubules. Between 40 and 45 days, the rete testis is predominantly an extratesticular rete situated in the cranial peduncle of the gonadal fold and in broad contact with the pro/mesonephric giant corpuscle. During this period, the intragonadal rete enters the gonad proper from its craniodorsal pole and extends into the cranial fourth of the testis. Between 60 and 110 days the rete testis attains its definitive position, extending into the central longitudinal axis as far as to the caudal fourth of the testis. For the caudal expansion of the rete testis the preceding proliferation of the mediastinal stroma is an important prerequisite. In the 40 to 45-day-old embryo the area of the testicular cords may be divided into two zones. A narrow outer zone contains plate-like cords with a thick diameter, and a larger central zone is filled with a network of thinner cords. Only the thick outer cords transform into the permanent seminiferous tubules, whereas the thinner cords in the central zone are transitory structures that disappear between 45 and 110 days. One important function of these transitory cords is to establish a continuous system of basal laminae that allows a direct connection between the central ends of the growing seminiferous tubules and the peripheral extensions of the rete testis (future straight testicular tubules). The first true straight testicular tubules become visible between 85 and 110 days. Due to a strong proliferation of the tubulus rectus-cells the straight testicular tubules elongate continuously, and the border between the rete system and the seminiferous tubules is slowly shifted towards the testicular periphery. This shift is not restricted to the prenatal period, but proceeds until after birth. At the cytological level, the formation and elongation of the straight testicular tubules is effected by proliferating cells that advance along the continuous basal lamina into the area of the seminiferous tubules. The pre-Sertoli and germ cells in this zone of invasion are separated from each other and overgrown by the tubulus rectus-cells. Exposed to the special milieu of the straight testicular tubules, pre-Sertoli and germ cells apparently cannot survive and finally disappear.

Prespermatogenesis and spermatogoniogenesis in the bovine testis.

The bovine male germ cell population was studied over the entire period from testicular differentiation in the embryo through onset of spermatogenesis in the pubertal calf. Germ cells were identified by protein gene product 9.5 immunohistochemistry and characterized by their ultrastructure. The proliferation pattern of germ cells was studied with immunohistochemical anti-Ki 67 and anti-proliferating cell nuclear antigen reactions. Germ cells with a high proliferation rate are observed from day 50 p.c. to day 80 p.c. These cells are in transition from primordial germ cells to prespermatogonia. From day 80 p.c. until approximately the 15th postnatal week the germ cells present are identified as prespermatogonia. From day 80 p.c. to day 200 p.c. germ cell multiplication decreases continuously; then the prespermatogonia enter a phase of relative mitotical quiescence that lasts until the 4th postnatal week. Between the 4th and the 15th postnatal week, testicular tubular diameters grow from 40 to 80 microm and the prespermatogonia resume their proliferation. In seminiferous tubules with diameters between 80 and 120 microm, found in animals between 18 and 27 weeks of age, a central lumen is normally still absent. During this period germ cell proliferation reaches a second maximum. The cells involved represent the members of the spermatogonia stem and precursor cell line kinetically interpolated between the prespermatogonia and the first differentiating A-spermatogonia. This second phase of prepubertal germ cell multiplication coincides with the period when the pre-Sertoli cells transform into adult-type Sertoli cells and enter the G0-phase for the rest of life.

The significance of rudimentary nephrostomial tubules for the origin of the vertebrate gonad.

Initial gonadal development was studied in 30- to 40-day-old bovine embryos. The results were interpreted in conjunction with findings on pro- and mesonephric organization in larval forms of Ichthyophis kohtaoensis (Gymnophiona, Amphibia). In bovine embryos vestigial nephrostomial tubules are the immediate precursors of the blastemas for adrenocortical, rete, gonadal and Mullerian infundibular development. From the study of Ichthyophis it can be concluded that the vestigial nephrostomial tubules seen in the bovine embryo are pronephric and not mesonephric in nature. As a consequence, the indifferent mammalian gonad is defined as a modified homologue of the pronephros situated in the zone of pro-/mesonephric overlapping. Such an overlapping of the two kidney generations in the fully developed state is clearly seen in Ichthyophis. Overlapping of the mesonephros with the modified pronephros (gonad) is necessary to allow intercalation of mesonephric tubules (efferent ductules in mammals) into the male seminal excurrent duct system.

Advanced glycosylation end products (AGEs), insulin-like growth factor-1 (IGF-1) and IGF-binding protein-3 (IGFBP-3) in patients with type 2 diabetes mellitus.

BACKGROUND: Advanced glycosylation end product (AGE) formation is a major mechanism for the development of complications in diabetes, and the possible roles of insulin-like growth factor 1 (IGF-1) and IGF binding protein 3 (IGFBP-3) are not clearly established. METHODS: We examined the associations of AGEs, free IGF-I and IGFBP-3 in Type 2 diabetes mellitus (DM) patients under diverse conditions. In a cross-sectional design we studied 110 subjects (67 women and 43 men): non-diabetic controls in group 1, (n = 15) and diabetes patients as follows: group 2, without complications (n = 25); group 3, with chronic complications (n = 25); group 4, with acute or chronic infections (n = 24); group 5, hospitalized for reasons unrelated to diabetes (n = 9); group 6, with end-stage renal disease (ESRD) (n = 12). AGEs were determined by a spectrofluorometric method (HPLC). Insulin and IGFBP-3 were measured by RIA and free IGF-1 with an IRMA method. RESULTS: AGEs were 13-fold higher in patients with ESRD (p<0.001), and lower in healthy individuals. Free IGF-1 was lower in the patients with complications (p = 0.017), with infections (p = 0.006) and hospitalized (p = 0.04). IGFBP-3 was higher in hospitalized patients (p=0.017). AGEs were associated with free IGF-1 (r = 0.41, p = 0.04) in the group with complications, and with HbA(1c) (r = -0.90, p = 0.002) in hospitalized patients. In the total group, free IGF-1 (r = -0.25, p = 0.008), and IGFBP-3 (r = -0.22, p = 0.021) were associated with HbA(1c). CONCLUSION: We concluded that AGEs were markedly increased in diabetic patients with ESRD, IGF-1 was decreased in patients with infections and hospitalized, and was negatively associated with HbA(1c). IGFBP-3 was increased in hospitalized patients, with higher levels in patients with long bone fractures. A complex interaction of humoral factors may participate in the acceleration of complications of diabetes.

The determination of 3-nitrophenol and some other aromatic impurities in 4-nitrophenol by reversed phase HPLC with peak suppression diode array detection.

In this work the peak suppression technique is used for the determination of 3-nitrophenol and some other aromatic impurities in 4-nitrophenol by reversed phase HPLC with diode array detection. Taking into account the differences between the absorption spectra of the two compounds, two wavelengths were selected in order to obtain the maximum difference between the spectral contribution for 3-nitrophenol and to maintain a small, similar spectral contribution for 4-nitrophenol (the main compound). Then we used the wavelength corresponding to a small spectral contribution of 3-nitrophenol as the reference wavelength. It was shown that taking lambda(an) = 266 nm and lambda(ref) = 364 nm, a broad elution peak of 4-nitrophenol was suppressed deconvoluting the peak of 3-nitrophenol. Moreover, quantitation of 3-nitrophenol was achieved without chemometric tools. Under the proposed conditions the detection limits for 3-nitrophenol and other common impurities of 4-nitrophenol used in the pharmaceutical industry (4-chlorophenol, 4-nitrophenol, 1-chloro-2-nitrobenzene, 1-chloro-4-nitrobenzene, 4,4'-bisfenilether, and 4,4'-dichloroazobenzene) were not significantly affected as compared with respective detection limits evaluated in the absence of 4-nitrophenol and using standard detection conditions (lambda(an) = 280 nm and lambda(ref) = 420 nm).

On the innervation of the donkey testis.

The innervation pattern of the adult donkey testis was investigated by immunohistochemistry and acetylcholinesterase histochemistry. Autonomous nerves reach the testis by three access-routes as funicular, mesorchial and caudal contributions. From these, the funicular contribution accompanying the testicular artery and pampiniform plexus is the strongest and most important one. Testicular innervation in the donkey is not uniform. The spermatic cord as well as the epididymal region, cranial and caudal poles (tunica albuginea and adjacent parenchyma and stroma) are well innervated, mostly by vascular nerves. Towards the free border of the testis, the nerve density in the tunica albuginea decreases continuously. In the interior of the gonad, approximately one third of the testis, situated between the free border and the central mediastinum, is practically devoid of any innervation. The great majority of the testicular nerves demonstrated by the present techniques are non-myelinated vascular nerves which react positive for dopamine-beta-hydroxylase and tyrosine hydroxylase, thus representing postjunctional sympathetic fibers. Many of these also contain neuropeptide Y. The testicular innervation of the donkey testis is free of cholinergic fibers. Calcitonin gene-related peptide-containing nerves are found as solitary varicose axons in the wall of blood vessels, but also in stromal connective tissue of the spermatic cord, tunica albuginea and septula testis.