Search

The CCR5-Δ32 mutation present in European populations is among the most prominently debated cases of recent positive selection in humans. This allele, a 32-bp deletion that renders the T-cell CCR5 receptor nonfunctional, has important epidemiological and public health significance, as homozygous carriers are resistant to several HIV strains. However, although the function of this allele in preventing HIV infection is now well described, its human evolutionary origin is poorly understood. Initial attempts to determine the emergence of the CCR5-Δ32 allele pointed to selection during the 14th-century Black Death pandemic; however, subsequent analyses suggest that the allele rose in frequency more than 5,000 years ago, possibly through drift. Recently, three studies have identified populations predating the 14th century CE that are positive for the CCR5-Δ32 allele, supporting the claim for a more ancient origin. However, these studies also suggest poorly understood regional differences in the recent evolutionary history of the CCR5-Δ32 allele. Here a new hydrolysis-probe-based real-time PCR assay was designed to ascertain CCR5 allele frequency in 53 individuals from a 10th- to 12th-century CE church and convent complex in central Germany that predates outbreaks of the Black Death pandemic. High-confidence genotypes were obtained for 32 individuals, and results show that CCR5-Δ32 allele frequency has remained unchanged in this region of Central Europe over the last millennium, suggesting that there has been no strong positive selective pressure over this time period and confirming a more ancient origin for the allele.

A role for GH and IGF-I in the modulation of the immune system has been under discussion for decades. Generally, GH is considered a stimulator of innate immune parameters in mammals and teleost fish. The stimulatory effects in humans as well as in bony fish often appear to be correlated with elevated endocrine IGF-I (liver-derived), which has also been shown to be suppressed during infection in some studies. Nevertheless, data are still fragmentary. Some studies point to an important role of GH and IGF-I particularly during immune organ development and constitution. Even less is known about the potential relevance of local (autocrine/paracrine) IGF-I within adult and developing immune organs, and the distinct localization of IGF-I in immune cells and tissues of mammals and fish has not been systematically defined. Thus far, IGF-I has been localized in different mammalian immune cell types, particularly macrophages and granulocytes, and in supporting cells, but not in T-lymphocytes. In the present study, we detected IGF-I in phagocytic cells isolated from rainbow trout head kidney and, in contrast to some findings in mammals, in T-cells of a channel catfish cell line. Thus, although numerous analogies among mammals and teleosts exist not only for the GH/IGF-system, but also for the immune system, there are differences that should be further investigated. For instance, it is unclear whether the primarily reported role of GH/IGF-I in the innate immune response is due to the lack of studies focusing on the adaptive immune system, or whether it truly preferentially concerns innate immune parameters. Infectious challenges in combination with GH/IGF-I manipulations are another important topic that has not been sufficiently addressed to date, particularly with respect to developmental and environmental influences on fish growth and health.

Many studies have been concerned with the ancient Egyptian mummification method; nevertheless, little effort has been made to explore it experimentally. The goal of this study is to apply evidence-based diagnostic criteria and state-of-the art methodology in order to improve knowledge on soft tissues preservation and postmortem alterations. Two human lower limbs (LL) from a female donor were (1) "naturally" mummified by dry heat and (2) artificially in natron. At specific time intervals a macroscopic and radiological examination of the LL was performed and skin and muscle samples were taken for histological and biomolecular analysis. Temperature, humidity, pH, and weight of the LL were systematically measured. The mummification by dry heat was stopped after 7 days due to unexpected lack of mummification progress. The mummification in natron was completed successfully after 208 days. The humidity, the external temperature, and the pH were proven with Pearson correlation and principal component analysis as important factors for the mummification process. The steady removal of water from the tissues through the natron has prevented the putrefaction. This is also evident in the absence of bacteria or fungi through the microbiological analysis. The histological analysis revealed very good preservation of the skin and the muscle tissues. In the muscular sample certain degree of structural disintegration can be seen, particularly affecting the epimysium whilst in the skin samples the epidermis, especially the stratum corneum, is mostly affected. The samples show better preservation compared with ancient Egyptian sections and other mummified tissues from historic or forensic context., (© 2015 Wiley Periodicals, Inc.)

Mummified human tissues are of great interest in forensics and biomolecular archaeology. The aim of this study was to analyse post mortem DNA alterations in soft tissues in order to improve our knowledge of the patterns of DNA degradation that occur during salt mummification. In this study, the lower limb of a female human donor was amputated within 24 h post mortem and mummified using a process designed to simulate the salt dehydration phase of natural or artificial mummification. Skin and skeletal muscle were sampled at multiple time points over a period of 322 days and subjected to genetic analysis. Patterns of genomic fragmentation, miscoding lesions, and overall DNA degradation in both nuclear and mitochondrial DNA was assessed by different methods: gel electrophoresis, multiplex comparative autosomal STR length amplification, cloning and sequence analysis, and PCR amplification of different fragment sizes using a damage sensitive recombinant polymerase. The study outcome reveals a very good level of DNA preservation in salt mummified tissues over the course of the experiment, with an overall slower rate of DNA fragmentation in skin compared to muscle.

Intensified aquaculture has strong impact on fish health by stress and infectious diseases and has stimulated the interest in the orchestration of cytokines and growth factors, particularly their influence by environmental factors, however, only scarce data are available on the GH/IGF-system, central physiological system for development and tissue shaping. Most recently, the capability of the host to cope with tissue damage has been postulated as critical for survival. Thus, the present study assessed the combined impacts of estrogens and bacterial infection on the insulin-like growth factors (IGF) and tumor-necrosis factor (TNF)-α. Juvenile rainbow trout were exposed to 2 different concentrations of 17β-estradiol (E2) and infected with Yersinia ruckeri. Gene expressions of IGF-I, IGF-II and TNF-α were measured in liver, head kidney and spleen and all 4 estrogen receptors (ERα1, ERα2, ERβ1 and ERβ2) known in rainbow trout were measured in liver. After 5 weeks of E2 treatment, hepatic up-regulation of ERα1 and ERα2, but down-regulation of ERß1 and ERß2 were observed in those groups receiving E2-enriched food. In liver, the results further indicate a suppressive effect of Yersinia-infection regardless of E2-treatment on day 3, but not of E2-treatment on IGF-I whilst TNF-α gene expression was not influenced by Yersinia-infection but was reduced after 5 weeks of E2-treatment. In spleen, the results show a stimulatory effect of Yersinia-infection, but not of E2-treatment on both, IGF-I and TNF-α gene expressions. In head kidney, E2 strongly suppressed both, IGF-I and TNF-α. To summarise, the treatment effects were tissue- and treatment-specific and point to a relevant role of IGF-I in infection., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Oestrogens and insulin-like growth factors (Igfs) play both a central role in the regulation of reproduction and growth and can interact especially in species showing a clear-cut sex-linked growth dimorphism (SGD) like in tilapia. Aromatase is essential in ovarian differentiation and oogenesis since it controls oestrogen synthesis. During tilapia sex differentiation, aromatase cyp19a1a expression increases from 9 days post-fertilization (dpf), resulting in high oestradiol level. High temperature, exogenous androgens or aromatase inhibitors override genetic sex differentiation inducing testes development through the suppression of cyp19a1a gene expression and aromatase activity. Supplementation with 17ß-oestradiol (E2) of gonadectomized juveniles induced a sustained and higher E2 plasma level than in intact or gonadectomized controls and both sexes showed reduced growth. Juvenile and mature females treated with the aromatase inhibitor 1,4,6-androstatriene-3,17-dione had 19% lower E2 plasma level compared to controls and they showed a 32% increased growth after 28 days of treatment. Altogether, these data suggest that E2 inhibits female growth leading to the SGD. Regarding Igf-1, mRNA and peptide appeared in liver at ∼ 4 dpf and then in organs involved in growth and metabolism, indicating a role in early growth, metabolism and organogenesis. Gonad igf-1 showed an early expression and the peptide could be detected at ∼ 7 dpf in somatic cells. It appeared in germ cells at the onset of ovarian (29 dpf) and testicular (52 dpf) meiosis. In testis, Igf-1 together with steroids may regulate spermatogenesis whereas in ovary it participates in steroidogenesis regulation. Igf-1 and Igf-2 promote proliferation of follicular cells and oocyte maturation. Igf-3 expression is gonad specific and localized in the ovarian granulosa or testicular interstitial cells. In developing gonads igf-3 is up-regulated in males but down-regulated in females. In contrast, bream Gh injections increased igf-1 mRNA in male and female liver and ovaries but gonadal igf-3 was not affected. Thus, local Igf-1 and Igf-2 may play crucial roles in the formation, development and function of gonads while Igf-3 depending on the species is involved in male and female reproduction. Furthermore, precocious ethynylestradiol (EE) exposure induced lasting effects on growth, through pituitary gh inhibition, local suppression of igf-1 expression and in testis only down-regulation of igf-3 mRNA. In conclusion, SGD in tilapia may be driven through an inhibitory effect due to E2 synthesis in female and involving Igfs regulation., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The examination was applied to sampling of 17 patients with stable implants after 18-36 months after endoprosthesis replacement of knee joint and to sampling of 17 patients with developed within this time-frame aseptic instability of endoprosthesis within the same time-frame. The typing of lymphocytes of peripheral blood was implemented using laser flow cytometry. The detection of immunoglobulins, cytokines, circulating immune complexes was made applying the technique of enzymoimmunoassay. In case of development of aseptic instability the signs testifying moderate activation of immune system and starting dysfunction of monocyte macrophage component of immune system. At that, IL-6, TNFα, CD14+HLA-DR provided the most complete information on the side of diagnostic of developing aseptic instability of joint.

Calcified dental plaque (dental calculus) preserves for millennia and entraps biomolecules from all domains of life and viruses. We report the first, to our knowledge, high-resolution taxonomic and protein functional characterization of the ancient oral microbiome and demonstrate that the oral cavity has long served as a reservoir for bacteria implicated in both local and systemic disease. We characterize (i) the ancient oral microbiome in a diseased state, (ii) 40 opportunistic pathogens, (iii) ancient human-associated putative antibiotic resistance genes, (iv) a genome reconstruction of the periodontal pathogen Tannerella forsythia, (v) 239 bacterial and 43 human proteins, allowing confirmation of a long-term association between host immune factors, 'red complex' pathogens and periodontal disease, and (vi) DNA sequences matching dietary sources. Directly datable and nearly ubiquitous, dental calculus permits the simultaneous investigation of pathogen activity, host immunity and diet, thereby extending direct investigation of common diseases into the human evolutionary past.

We present a comparative analysis of the allelic polymorphism of the matrix metalloproteinase (MMP) gene family, including MMP3 (rs3025058), MMP7 (rs11568818), MMP9 (rs17576, rs2250889), MMP12 (rs2276109), and MMP13 (rs2252070), in patients with external genital endometriosis (EGE) and in a control group of healthy women proven to be free of disease by laparoscopic inspection. We found significant differences in the incidence of particular MMP3 and MMP9 alleles, which substantiate the role of matrix metalloproteinases in EGE pathogenesis. We used the Multifactor Dimensionality Reduction (MDR) analysis to show that 14 allelic combinations of the MMP containing MMP3 (rs3025058) x MMP7 (rs11568818) x MMP9 (rs17576) alleles showed a statistically significant association with an increased risk of EGE, while 10 other combinations correlated with a reduced risk of the disease. MDR analysis produced two statistically significant models for MMP allelic combinations involved in EGE progression, both with 100% penetrance and 83% and 78% accuracy.

Jörg Jenatsch was a Swiss defender of independence and a fighter for liberty in the 17th century. With the help of three living male members of the Jenatsch family, we successfully identified a skeleton exhumed from Chur cathedral as the remains of Jörg Jenatsch. Our conclusion was based upon complete Y-STR and Y-SNP profiles that could be generated by replicate analyses of a bone sample available to us. The skeleton and the three living family members carried the same Y-SNP haplogroup, but were discordant at three of 23 Y-STR loci. This notwithstanding, conservative biostatistical evaluation of the data suggests that the Chur skeleton is at least 20 times more likely than not to be Jörg Jenatsch., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Objective: To assess changes in different tissues during the process of artificial mummification by natron using computed tomography (CT) and magnetic resonance imaging (MRI), and to translate the results to image interpretation in paleoradiological studies of ancient mummies., Materials and Methods: A human lower limb (LL) was amputated from a female donor 24 h post-mortem and mummified by artificial natron (54 % NaCl, 16 % Na2SO4, 18 % Na2CO3 12 % NaHCO3) in ancient Egyptian style. The LL was kept in a fume hood at 16-25 °C and 30-75 % relative humidity. CT and MRI were performed at specific intervals with quantitative evaluation of Hounsfield units (HU) and signal intensities (SI)., Results: Evaluated tissues showed different HU and SI changes during the experimental mummification. All tissues revealed an overall but varying increase of HU in CT examinations. All tissues except for the compact bone revealed an overall but varying decrease of SI in the IR and T2-weighted sequences of the MRI. Typical findings included a distinct increase of HU in the cutis at the end of the study and a temporary increase of SI in the IR and T2-weighted sequences in all muscle groups., Conclusions: Radiological findings showed a regular, controlled and effective dehydration by the applied natron without detectable putrefaction. Evaluated tissues revealed different radiological changes during the experiment, which altogether led to preservation of the tissues without radiologically identifiable destruction. The cutis revealed radiological signs of direct interaction with the natron in the form of covering and possibly permeation.

Aim: To investigate the effect of local intestinal perfusion with hypertonic saline (HTS) on intestinal ischemia-reperfusion injury (IRI) in both ex vivo and in vivo rat models., Methods: All experiments were performed on male Wistar rats anesthetized with pentobarbital sodium given intraperitoneally at a dose of 60 mg/kg. Ex vivo vascularly perfused rat intestine was subjected to 60-min ischemia and either 30-min reperfusion with isotonic buffer (controls), or 5 min with HTS of 365 or 415 mOsm/L osmolarity (HTS(365mOsm) or HTS(415mOsm), respectively) followed by 25-min reperfusion with isotonic buffer. The vascular intestinal perfusate flow (IPF) rate was determined by collection of the effluent from the portal vein in a calibrated tube. Spontaneous intestinal contraction rate was monitored throughout. Irreversible intestinal injury or area of necrosis (AN) was evaluated histochemically using 2.3.5-triphenyltetrazolium chloride staining. In vivo, 30-min ischemia was followed by either 30-min blood perfusion or 5-min reperfusion with HTS(365mOsm) through the superior mesenteric artery (SMA) followed by 25-min blood perfusion. Arterial blood pressure (BP) was measured in the common carotid artery using a miniature pressure transducer. Histological injury was evaluated in both preparations using the Chui score., Results: Ex vivo, intestinal IRI resulted in a reduction in the IPF rate during reperfusion (P < 0.05 vs sham). The postischemic recovery of the IPF rate did not differ between the controls and the HTS(365mOsm) group. In the HTS(415mOsm) group, postischemic IPF rates were lower than in the controls and the HTS(365mOsm) group (P < 0.05). The intestinal contraction rate was similar at baseline in all groups. An increase in this parameter was observed during the first 10 min of reperfusion in the control group as compared to the sham-treated group, but no such increase was seen in the HTS(365mOsm) group. In controls, AN averaged 14.8% ± 5.07% of the total tissue volume. Administration of HTS(365mOsm) for 5 min after 60-min ischemia resulted in decrease in AN (5.1% ± 1.20% vs controls, P < 0.01). However, perfusion of the intestine with the HTS of greater osmolarity (HTS(415mOsm)) failed to protect the intestine from irreversible injury. The Chiu score was lower in the HTS(365mOsm) group in comparison with controls (2.4 ± 0.54 vs 3.2 ± 0.44, P = 0.042), while intestinal perfusion with HTS(415mOsm) failed to improve the Chiu score. Intestinal reperfusion with HTS(365mOsm) in the in vivo series secured rapid recovery of BP after its transient fall, whereas in the controls no recovery was seen. The Chiu score was lower in the HTS(365mOsm) group vs controls (3.1 ± 0.26 and 3.8 ± 0.22, P = 0.0079 respectively,), although the magnitude of the effect was lower than in the ex vivo series., Conclusion: Brief intestinal postischemic perfusion with HTS(365mOsm) through the SMA followed by blood flow restoration is a protective procedure that could be used for the prevention of intestinal IRI.

Vitellogenin (Vg) of the barfin plaice Liopsetta pinnifasciata was isolated and purified. In native polyacrylamide gel electrophoresis, Vg appeared as one band. After being subjected to sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), Vg fraction produced several polypeptides with molecular masses of 180, 98, 70, 52, 41 and 37 kDa. MALDI-TOF mass spectrometry (MS) of the 180- and 98-kDa Vg polypeptides from the SDS-PAGE gel and de novo sequencing of their four peptide fragments based on MS/MS analysis confirmed that the purified proteins were vitellogenins, which shared high similarity with the Vgs of the barfin flounder Verasper moseri and Atlantic halibut Hippoglossus hippoglossus. The most part of the predicted sequences obtained from the L. pinnifasciata 180-kDa polypeptide has previously been found in the V. moseri vitellogenin type B, the sequences obtained from the 98-kDa polypeptide were found in V. moseri vitellogenin type A, so these findings allow us to propose that L. pinnifasciata has at least two different forms of Vg. Rabbit polyclonal antibodies against Vg were produced, and a quantitative enzyme-linked immunosorbent assay was developed. The concentration of Vg in barfin plaice from the moderately contaminated area of Amursky Bay in the Sea of Japan was detected based on the maturity stage of their gonads. In November 2008, the Vg concentration in the plasma of females with advanced oogenesis varied from 5.295 to 28.367 mg/ml (mean 16.38 ± 6.73 mg/ml, CV = 41.1%); in the plasma of males, the concentration ranged from non-detectable to 0.957 mg/ml (0.29 ± 0.42 mg/ml, CV = 127.9%). In October 2009, the Vg concentration in female plasma was lower than in November 2008 (2.21-13.87 mg/ml). High individual variability of plasma Vg was characteristic for maturing males (CV = 200.3%) and immature females (CV = 255.5%), and there was no significant difference between plasma Vg concentrations in males captured in November 2008 and October 2009 or in maturing males and immature females. Vacuolisation of hepatocytes was more typical for males with low plasma Vg concentrations and females with high plasma Vg concentrations. Necrosis and pyknosis of hepatocyte nuclei were more frequent in males with high Vg concentrations and in females with low plasma Vg concentrations.

Contradictory studies suggest IGF-I in fish liver and gills is involved in osmoregulation, but nothing is known about the kidney and intestine's role nor about IGF-II's role in any organ. Tilapia were transferred from freshwater (FW) to seawater (SW) for 1week (wk) and retransferred to FW for another week. At 4h, 1d, 2d, 3d and 1wk after SW-transfer and FW-retransfer IGF-I, IGF-II and growth hormone receptor (GHR1) mRNA were measured by real-time PCR. Hepatic IGF-I, IGF-II and GHR1 mRNA were downregulated in parallel after SW-transfer, recovered and were again downregulated after FW-retransfer. In gills, IGF-I, IGF-II and GHR1 were upregulated synchronously after SW-transfer and, partially also after FW-retransfer. The renal genes were downregulated after SW-transfer and partially upregulated after FW-retransfer. Persisting upregulation in intestinal IGF-I mRNA occurred after FW-retransfer. Thus, endocrine and auto/paracrine IGF-I and IGF-II seem to be involved in fish osmoregulation in an organ-specific manner., (Copyright 2010 Elsevier Ireland Ltd. All rights reserved.)

Insulin-like growth factor (IGF)-I, which is crucially involved in fish growth, differentiation, and reproduction, occurs in tilapia pituitary. IGF-I peptide, which is probably produced in hypothalamic perikarya, is present in axons of the neurohypophysis, and IGF-I mRNA and peptide are present in the adenohypophysis in adrenocorticotrophic hormone cells, melanocyte-stimulating hormone cells, with interindividual differences in growth hormone cells, and, starting with puberty, in gonadotrophic hormone (GTH) cells. Subordinate males showed a high IGF-I but a lower beta-luteinizing hormone expression, while in dominant males the opposite was found. IGF-I from the GTH cells may act as auto/paracrine regulators of GTH cell proliferation and enhance GTH synthesis and release during puberty and reproduction, depending on the social status.

The enormous expansion of world-wide aquaculture has led to increasing interest in the regulation of fish immune system. Estrogen has recently been shown to inhibit the endocrine (liver-derived) and autocrine/paracrine local insulin-like growth factor-I system in fish. In order to address the potential actions of estrogen on the IGF system in immune organs, tilapia were fed with 17alpha-ethinylestradiol (EE2)-enriched food from 10 to 40 days post fertilization (DPF) to induce functional feminization, an approach commonly used in aquaculture. EE2-treated and control fish were sampled at 75 and 165 DPF. The expression levels of ER-alpha, IGF-I, IGF-II and growth hormone receptor (GH-R) mRNA in spleen and head kidney were determined by real-time PCR and the expressing sites of IGF-I mRNA identified by in situ hybridisation. Ratios of spleen length and weight to body length and weight were determined. At 165 DPF, the length (4.9% vs. 7.6%) and weight (0.084% vs. 0.132%) ratios were significantly lowered in EE2-treated fish and number and size of the melanomacrophage centres were considerably reduced. At 75 DPF, both in spleen and head kidney of EE2-treated fish the expression levels of IGF-I and IGF-II mRNA were markedly diminished. The suppression was more pronounced for IGF-I (spleen: -12.071-fold; head kidney: -8.413-fold) than for IGF-II (spleen: -4.102-fold; head kidney: -1.342-fold). In agreement, clearly fewer leucocytes and macrophages in head kidney and spleen of EE2-treated fish contained IGF-I mRNA as shown by in situ hybridisation. ER-alpha mRNA expression in spleen was increased at 75 DPF but unchanged in head kidney. GH-R gene expression showed a mild upregulation at 165 DPF in both tissues. Thus, exposure to EE2 during early development affected distinctly the IGF system in tilapia immune organs. It led to lasting impairment of spleen growth and differentiation that can be attributed to an interaction of EE2 with IGF-I and, less pronouncedly, IGF-II. Especially, the impairment of spleen and melanomacrophage centres might interfere with the antigen presentation capacity of the immune system and, thus, alter susceptibility to infection.

The aim of this study was to evaluate whether effects of environmental estrogens on fish growth and reproduction may be mediated via modulating the growth hormone (GH)/insulin-like growth factor I (IGF-I) system. To this end, developing male and female monosex populations of tilapia were exposed to 17alpha-ethinylestradiol (EE2) at 5 and 25 ng EE2/l water from 10-day postfertilization (DPF) until 100 DPF. Under exposure to both EE2 concentrations, sex ratio shifted toward more females and body length, and weight were significantly reduced in males. The growth-reducing effect was associated with significant changes in hepatic IGF-I expression, both in males and females and with significant alterations of IGF-I mRNA and GH mRNA in the brain. The changes in IGF-I and GH mRNA were accompanied by altered estrogen receptor alpha (ERalpha) expression in brain and liver. These findings point to an influence of estrogenic exposure on the endocrine GH/IGF-I axis. In addition, the EE2 treatment resulted in significant changes of ERalpha and IGF-I expression in ovaries and testis, suggesting that the estrogens interact not only with the endocrine but also with the autocrine/paracrine part of the IGF-I system. Overall, our results provide evidence that EE2 at environmentally relevant concentrations is able to interfere with the GH/IGF-I system in bony fish and that the impairing effects of estrogens reported on fish growth and reproductive functions may rather result from a cross talk between the sex steroid and the IGF-I system than be toxicological.

Growth and sexual development are closely interlinked in fish; however, no reports exist on potential effects of estrogen on the GH/IGF-I-axis in developing fish. We investigate whether estrogen exposure during early development affects growth and the IGF-I system, both at the systemic and tissue level. Tilapia were fed from 10 to 40 days post fertilization (DPF) with 17alpha-ethinylestradiol (EE(2)). At 50, 75, 90, and 165 DPF, length, weight, sex ratio, serum IGF-I (RIA), pituitary GH mRNA and IGF-I, and estrogen receptor alpha (ERalpha) mRNA in liver, gonads, brain, and gills (real-time PCR) were determined and the results correlated to those of in situ hybridization for IGF-I. Developmental exposure to EE(2) had persistent effects on sex ratio and growth. Serum IGF-I, hepatic IGF-I mRNA, and the number of IGF-I mRNA-containing hepatocytes were significantly decreased at 75 DPF, while liver ERalpha mRNA was significantly induced. At 75 DPF, a transient decline of IGF-I mRNA and a largely reduced number of IGF-I mRNA-containing neurons were observed in the female brain. In both sexes, pituitary GH mRNA was significantly suppressed. A transient downregulation of IGF-I mRNA occurred in ovaries (75 DPF) and testes (90 DPF). In agreement, in situ hybridization revealed less IGF-I mRNA signals in granulosa and germ cells. Our results show for the first time that developmental estrogen treatment impairs GH/IGF-I expression in fish, and that the effects persist. These long-lasting effects both seem to be exerted indirectly via inhibition of pituitary GH and directly by suppression of local IGF-I in organ-specific cells.

Several lines of growth hormone (GH)-overexpressing fish have been produced and analysed for growth and fertility parameters. However, only few data are available on the growth-promoting hormone insulin-like growth factor I (IGF-I) that mediates most effects of GH, and these are contradictory. Using quantitative real-time RT-PCR, radioimmunoassay, in situ hybridization, immunohistochemistry, and radiochromatography we investigated IGF-I and IGF binding proteins (IGFBPs) in an adult (17 months old) transgenic (GH-overexpressing) tilapia (Oreochromis niloticus). The transgenics showed an around 1.5-fold increase in length and an approximately 2.3-fold higher weight than the non-transgenics. Using radioimmunoassay, the serum IGF-I levels were lower (6.22 +/- 0.75 ng/ml) in transgenic than in wild-type (15.01 +/- 1.49 ng/ml) individuals (P = 0.0012). Radioimmunoassayable IGF-I in transgenic liver was 4.2-times higher than in wild-type (16.0 +/- 2.21 vs. 3.83 +/- 0.71 ng/g, P = 0.0017). No hepatocytes in wild-type but numerous hepatocytes in transgenic liver contained IGF-I-immunoreactivity. RT-PCR revealed a 1.4-times higher IGF-I mRNA expression in the liver of the transgenics (10.51 +/- 0.82 vs. 7.3 +/- 0.49 pg/microg total RNA, P = 0.0032). In correspondence, in situ hybridization showed more IGF-I mRNA containing hepatocytes in the transgenics. A twofold elevated IGF-I mRNA expression was determined in the skeletal muscle of transgenics (0.33 +/- 0.02 vs. 0.16 +/- 0.01 pg/microg total RNA, P < 0.0001). Both liver and serum of transgenics showed increased IGF-I binding. The increased IGFBP content in the liver may lead to retention of IGF-I, and/or the release of IGF-I into the circulation may be slower resulting in accumulation of IGF-I in the hepatocytes. Our results indicate that the enhanced growth of the transgenics likely is due to enhanced autocrine/paracrine action of IGF-I in extrahepatic sites, as shown here for skeletal muscle.

In bony fish, IGF-I released from the liver under the control of pituitary GH is the main endocrine regulator of growth, maintenance and development, and the amount of circulating IGF-I regulates synthesis and release of GH. In mammals and amphibia, evidence indicates that anterior pituitary endocrine cells also contain IGF-I. However, only preliminary and conflicting data exist on IGF-I gene expression in bony fish pituitary. Thus, we investigated the presence of IGF-I in the tilapia (Oreochromis niloticus) pituitary by quantitative real-time RT-PCR, in situ hybridisation and immunohistochemistry. The absolute amount of IGF-I mRNA in the whole pituitary (7.4+/-3.3 x 10(-3)pg/microg total RNA) was 1000-times lower than in liver (7.5+/-3.1 pg/microg total RNA). IGF-I peptide occurred in both neuro- and adenohypophysis but IGF-I gene expression was mainly restricted to the adenohypophysis. In the neurohypophysis, only few cells, probably pituicytes, contained IGF-I mRNA whereas IGF-I peptide was found also in numerous axons in the pars nervosa. In the adenohypophysis, both IGF-I mRNA and peptide were present in the majority of ACTH cells in all individuals investigated. In alpha-MSH cells, only IGF-I mRNA but no IGF-I peptide was detected likely suggesting an immediate release of IGF-I after synthesis. IGF-I mRNA and peptide were further observed in GH cells but their presence showed pronounced inter-individual differences likely due to the physiological, e.g., nutritional, status of the individual. IGF-I released from the GH cells may serve as auto/paracrine mediator of a negative feedback mechanism in addition to liver-derived endocrine IGF-I. Generally, the constitutive synthesis of IGF-I in ACTH cells and the varying content in GH and alpha-MSH cells suggest particular roles for IGF-I. Local IGF-I may regulate synthesis and release of pituitary hormones in an autocrine and/or paracrine manner as well as prevent apoptosis and stimulate proliferation of endocrine cells.

Fish pituitary plays a central role in the control of growth, development, reproduction and adaptation to the environment. Several types of hormone-secreting adenohypophyseal cells have been characterised and localised in diverse teleost species. The results suggest a similar distribution pattern among the species investigated. However, most studies deal with a single hormone or hormone family. Thus, we studied adjacent sections of the pituitary of Oreochromis niloticus, the tilapia, by conventional staining and immunohistochemistry with specific antisera directed against growth hormone (GH), prolactin (PRL), somatolactin (SL), thyrotropin (beta-TSH), follicle-stimulating hormone (beta-FSH), luteinising hormone (beta-LH), adrenocorticotropic hormone (ACTH) and melanocyte-stimulating hormone (alpha-MSH). The pituitary was characterised by a close interdigitating neighbourhood of neurohypophysis (PN) and adenohypophysis. PRL-immunoreactive and ACTH-immunoreactive cells were detected in the rostral pars distalis. GH-immunoreactive cells were present in the proximal pars distalis (PPD). A small region of the PPD contained beta-TSH-immunoreactive cells, and beta-LH-immunoreactive cells covered approximately the remaining parts. Centrally, beta-FSH-immunoreactive cells were detected in the vicinity of the GH-containing cells. Some of these cells also displayed beta-LH immunoreactivity. The pars intermedia was characterised by branches of the PN surrounded by SL-containing and alpha-MSH-immunoreactive cells. The ACTH and alpha-MSH antisera were observed to cross-react with the respective antigens. This cross-reactivity was abolished by pre-absorption. We present a complete map of the distinct localisation sites for the classical pituitary hormones, thereby providing a solid basis for future research on teleost pituitary.

The cellular sites of insulin-like growth factor I (IGF-I) synthesis in the early developing tilapia (0-140 days post fertilization, DPF) were investigated. IGF-I mRNA and peptide appeared in liver as early as 4 DPF and in gastro-intestinal epithelial cells between 5-9 DPF. In exocrine pancreas, the expression of IGF-I started at 4 DPF and continued until 90 DPF. IGF-I production was detected in islets at 6 DPF in non-insulin cells and occurred throughout life. In renal tubules and ducts, IGF-I production started at 8 DPF. IGF-I production in chondrocytes had its onset at 4 DPF, was more pronounced in growing regions and was also found in adults. IGF-I mRNA and peptide appeared in the cytoplasm of skeletal muscle cells at 4 DPF. In gill chloride cells, IGF-I production started at 6 DPF. At 13 DPF, IGF-I was detected in cardiac myocytes. IGF-I-producing epidermal cells appeared at 5 DPF. In brain and ganglia, IGF-I was expressed in virtually all neurones from 6 to 29 DPF, their number decreasing with age. Neurosecretory IGF-I-immunoreactive axons were first seen in the neurohypophysis around 17 DPF. Endocrine cells of the adenohypophysis exhibited IGF-I mRNA at 28 DPF and IGF-I immunoreactivity at 40 DPF. Thus, IGF-I appeared early (4-5 DPF), first in liver, the main source of endocrine IGF-I, and then in organs involved in growth or metabolism. The expression of IGF-I was more pronounced during development than in juvenile and adult life. Local IGF-I therefore seems to have a high functional impact in early growth, metabolism and organogenesis.

The present study investigates the expression of IGF-I in the early developing tilapia (Oreochromis niloticus). IGF-I was detected very early in ontogeny (4-5 days postfertilization, DPF), first in liver and in organs involved in growth and metabolism, thus suggesting a high physiological impact of IGF-I in growth, metabolism, and organogenesis.

Polymorphysms of the three genes encoding phase 1 (CYP1A1, mEPH1, and CYP2E2) and the three genes encoding phase 2 (NAT2, GSTM1, and GSTT1) xenobiotic detoxication enzymes were typed by use of PCR in 74 patients with extragenital endometriosis. Distribution of the CYP1A1, mEPHX1, CYP2E1, NAT2, and GSTM1 polymorphic alleles in the patient group corresponded to that in the control group. At the same time, functionally defective genotypes GSTM1 0/0, NAT2 S/S; GSTM1 0/0, GSTT1 0/0; and GSTT1 0/0, NAT2 S/S were three, four and eight times more frequent among the patients than in healthy individuals. This observation suggests the existence of a distinct association between the functionally defective alleles of the phase 2 xenobiotic detoxication and endometriosis. Possible mechanisms underlying this association are discussed. It is suggested that typing of the NAT2, GSTM1, and GSTT1 genes can be useful for the assessment of the predisposition to endometriosis.

The relative frequencies of the normal (+) and null (0) alleles of the glutathione-S-transferase M1 (GSTM1) gene, as well as those of the rapid (R) and slow (S) forms of N-acetyl transferase 2 (NAT-2), were studied in the Russian and French populations and in endometriosis (EM) patients. In the total Russian and French populations, the proportions of homozygotes for deletion in gene GstM1 (0/0) were 42.2 and 45.8%, respectively, whereas in Russian and French EM patients, these values were 58.6 and 76.9%, respectively. The differences in these proportions between the total population and subjects with EM were significant at the confidence levels of 0.98 (chi 2 = 5.45; P < 0.02) and 0.90 (chi 2 = 3.01; P < 0.1) for the French and Russian populations, respectively. The frequencies of allele S of the Nat-2 gene were also similar in the Russian and French populations (60 and 63.1%, respectively), with these frequencies being somewhat higher in EM patients (71.2 and 77.7%, respectively). In Russians, the proportion of EM patients who were homozygous for the R form of NAT-2 (R/R) was significantly lower (chi 2 = 5.1). Forty-three of the patients with external genital EM received complex treatment with the use of the interferon inducer Cyclopheron. In 17 patients, a pronounced positive dynamics was observed, and 29 patients exhibited an increased resistance to the immunomodulating therapy. These groups comprised 1 and 25 GstM1 0/0 homozygotes, respectively; the number of patients with the slow NAT-2 form was 13 (7 S/S and 6 S/R genotypes) and 29 (20 S/S and 9 S/R genotypes), respectively. The obtained data indicate that the GstM1 and Nat-2 genes are involved in the EM pathogenesis. Therefore, molecular screening for the GstM1 0 and Nat-2 S alleles would be a good prognostic test when prescribing the postoperative treatment for EM and predicting its effectiveness.

Immunofluorescence was used to study the count of joint synovial membrane cells producing immunoglobulins of major classes (A, M, G) in 30 rabbits on a model of experimental arthrosis and the effects of rumalone, rumalone + indomethacin, and food deprivation. Clinically, the levels of immune responses in patients with primary osteoarthrosis were judged from the content of rumalone-sensitive serum antibodies. The development of experimental arthrosis was found to be followed by a substantial increase in the count of synovial membrane cells locally elaborating immunoglobulins chiefly G and M. With this, the use of pathogenetically substantiated therapy with chondroprotective agents produced a slight effect on the degree of local immune response impairments. Correction of the latter may be made with indomethacin and food deprivation in the experiment, as well as with indomethacin and microwave resonance therapy in the clinical setting.

Comparative evaluation of the efficacy of stimulating and hypobiotic agents in 122 dogs with experimentally induced cardiogenic shock and 56 patients with acute myocardial infarction complicated by cardiogenic shock allowed a relationship to be established of the efficiency of therapeutic measures to the phase and rate of progression of the pathological process. In rapid progression of clinical symptoms of cardiogenic shock it is predominantly preparations of hypobiotic action that are to be prescribed whereas in slow progression of these it is advisable to use agents of stimulating action.

A study is presented of the thermal semeiotics and the state of cellular and humoral immunity in 29 patients with osteoarthrosis deformans, 35 patients with rheumatoid arthritis of different activity. It is recommended to use data of the thermographic picture and immunological status for objectivation of the character of the pathological process, differential diagnosis and dynamic express-evaluation of treatment adequacy.

In 54 patients with osteoarthrosis deformans traditional treatment was supplemented by an antioxidant agent emoxipin. The drug inhibited lipid peroxidation, activated the antioxidant defense system with simultaneous regression of clinical manifestations. Results of the investigation evidence the efficiency of emoxipin in osteoarthrosis deformans.

Samples of chorionic villi and embryonic tissues (brain, brain--sheaths) are thoroughly washed with Hank's solution, immediately subjected to hypotonic treatment (0.9% sodium citrate plus few drops of 0.01% colchicine) 37 degrees C, 30 min, prefixed 20 min with equal amount of standard fixative mixture, twice fixed in standard fixative solution (1 hour, -10 degrees C), hydrated with equal volume of distilled water (5-10 min), dried, macerated directly on the slide with 60% acetic acid. The cell suspension is then evenly spread on the slide surface, dried, postfixed and stained. The method provides sufficient amount of metaphase and prometaphase mitotic plates suitable for differentiating staining in 1.5-2 hours after sampling and might be recommended for routine chromosomal analysis in prenatal diagnosis of inherited diseases during early pregnancy.

Using original direct methods of shaking-imprinting and rapid sampling of metaphase and prometaphase chromosomes, a cytogenetic analysis has been performed in 80 diagnostic chorionic villus samples (CVS) in the first trimester. Chromosomal disorders were identified in 6 (7.5%) fetuses: Down's syndrome in 4, monosomy 18 in 1, and a short-arm deletion of chromosome 18 in 1. These pregnancies were terminated. The paper offers opportunities for reducing risks of transcervical CVS and widely applying first-trimester chromosomal analysis in a program of mass screening for chromosomal and genetic diseases.

A study of 64 patients with acute myocardial infarction revealed an elevated concentration of peroxidation radicals in the blood, a reduction of the antioxidant system. Three hours after i/v administration of riboxin the content of peroxidation radicals reduced. After discontinuation of intravenous riboxin peroxidation of lipids increased again.