The influence of cryopreservation and quick-freezing on the mechanical properties of tendons

In order to maintain their native properties, cryopreserved tendons are usually used in biomechanical research and in transplantation of allogenic tendon grafts. The use of different study protocols leads to controversy in literature and thus complicates the evaluation of the current literature. The aim of this study consisted in examining the influence of different freezing and thawing temperatures on the mechanical properties of tendons. 60 porcine tendons were frozen at either −80 °C or −20 °C for 7 days and thawed at room or body temperature for 240 or 30 min, respectively. A subgroup of ten tendons was quick-frozen with liquid nitrogen (−196 °C) for 2 s before cryopreservation. Biomechanical testing was performed with a material testing machine and included creep, cyclic and load-to-failure tests. The results showed that freezing leads to a reduced creep strain after constant loading and to an increased secant modulus. Freezing temperature of −80 °C increased the secant modulus and decreased the strain at maximum stress, whereas thawing at room temperature reduced the maximum stress, the strain at initial tendon failure and the Young’s Modulus. Quick-freezing led to increased creep strain after constant loading, increased strain at initial failure in the load-to-failure test, and decreased strain at maximum stress. When cryopreserving, tendons for scientific or medical reasons, freezing temperature of −20 °C and thawing temperature of 37.5 °C are recommended to maintain the native properties of tendons. A treatment with liquid nitrogen in the sterilization process of tendon allografts is inadvisable because it alters the tendon properties negatively.     

乌梢蛇输精管道的组织学与免疫细胞化学观察

为探讨乌梢蛇(Zaocys dhumnades)输精管道结构与其功能之间的关系,该研究用一般光镜技术观察了乌梢蛇输精管道的显微结构及其年周期变化,并结合免疫细胞化学方法研究了雄激素受体(AR)、雌激素受体(ER)、孕激素受体(PR)和芳香化酶(Ar)在输精管和精巢中精子细胞表达的相关性.为验证该文在乌梢蛇输精管中观察到的大量精子和圆球状结构,用一般光镜技术还观察了黑眉锦蛇(Elaphe taeniura)、赤链蛇(Dinodon rufozonatum)与虎斑颈槽蛇(Rhabdophis tigrina lateralis)的输精管道.结果表明,乌梢蛇的输精管道主要由输出小管、附睾管与输精管构成;8-10月输出小管中有精子,8月—翌年1月附睾管中有精子,全年(除7月外)输精管中有大量精子;在输精管内首次观察到由多个精子细胞构成的圆球状结构,该结构与精巢中精子细胞的AR、ER、PR和Ar累计光密度值之间分别无显著差异.由于在乌梢蛇、黑眉锦蛇及赤链蛇的输精管内圆球状结构均可见精子细胞变态形成精子.因此,建议将蛇类输精管内圆球状结构命名为生精小球(seminiferous spherule).该文认为,蛇类精巢是精子形成的主要部位,而输精管内的生精小球是精子形成的另一个部位;附睾与输精管均可以储存精子,但输精管是精子储存的主要器官.     

似鲇高原鳅皮肤与鳍的组织学观察

采用石蜡切片技术及H.E和AB-PAS染色方法,对似鲇高原鳅(Triplophysa siluroides)头部、腹部、背部、侧线部和尾部皮肤结构及胸鳍、腹鳍、背鳍、臀鳍和尾鳍的组织结构进行观察。各部位皮肤均由表皮和真皮构成,真皮包括疏松层和致密层,不同部位皮肤厚度不同。表皮层腹部最厚,为(84.62±10.82)μm,侧线部最薄,为(14.97±3.95)μm,各部位表皮厚度差异显著。表皮层分布着黏液细胞、棒状细胞及味蕾。疏松层头部最厚,为(282.71±70.56)μm,尾部最薄,为(29.07±4.88)μm,该层分布有黑色素细胞、空泡状细胞和颗粒腺,而黏液腺分布于致密层。各部位的鳍均由表皮层、胶原纤维层、胶原下层及鳍条构成,表皮层与皮肤表皮层组成相似,鳍条是矿化的结缔组织。     

Crocin improved locomotor function and mechanical behavior in the rat model of contused spinal cord injury through decreasing calcitonin gene related peptide (CGRP)

Various approaches have been offered to alleviate chronic pain resulting from spinal cord injuries (SCIs). Application of herbs and natural products, with potentially lower adverse effects, to cure diseases has been recommended in both traditional and modern medicines. Here, the effect of crocin on chronic pain induced by spinal cord contusion was investigated in an animal model. Female Wistar rats were randomly divided into five groups (5 rats in each); three groups were contused at the L1 level. One group was treated with crocin (150 mg/kg) two weeks after spinal cord injury; the second group, control, was treated with vehicle only; and the third group was treated with ketoprofen. Two normal groups were also considered with or without crocin treatment. The mechanical behavioral test, the locomotor recovery test and the thermal behavioral test were applied weekly to evaluate the injury and recovery of rats. Significant improvements (p < 0.05) in mechanical behavioral and locomotor recovery tests were seen in the rats treated with crocin. Thermal behavioral test did not show any significant changes due to crocin treatment. Plasma concentration of calcitonin-gene related peptide (CGRP) changed from 780.2 ± 2.3 to 1140.3 ± 4.5 pg/ml due to SCI and reached 789.1 ± 2.7 pg/ml after crocin treatment. These changes were significant at the level of p < 0.05. The present study shows the beneficial effects of crocin treatment on chronic pain induced by SCI, through decreasing CGRP as an important mediator of inflammation and pain.     

The mechanical environment of the chondrocyte: a biphasic finite element model of cell-matrix interactions in articular cartilage

Mechanical compression of the cartilage extracellular matrix has a significant effect on the metabolic activity of the chondrocytes. However, the relationship between the stress–strain and fluid-flow fields at the macroscopic “tissue” level and those at the microscopic “cellular” level are not fully understood. Based on the existing experimental data on the deformation behavior and biomechanical properties of articular cartilage and chondrocytes, a multi-scale biphasic finite element model was developed of the chondrocyte as a spheroidal inclusion embedded within the extracellular matrix of a cartilage explant. The mechanical environment at the cellular level was found to be time-varying and inhomogeneous, and the large difference (3 orders of magnitude) in the elastic properties of the chondrocyte and those of the extracellular matrix results in stress concentrations at the cell–matrix border and a nearly two-fold increase in strain and dilatation (volume change) at the cellular level, as compared to the macroscopic level. The presence of a narrow “pericellular matrix” with different properties than that of the chondrocyte or extracellular matrix significantly altered the principal stress and strain magnitudes within the chondrocyte, suggesting a functional biomechanical role for the pericellular matrix. These findings suggest that even under simple compressive loading conditions, chondrocytes are subjected to a complex local mechanical environment consisting of tension, compression, shear, and fluid pressure. Knowledge of the local stress and strain fields in the extracellular matrix is an important step in the interpretation of studies of mechanical signal transduction in cartilage explant culture models.     

Evaluation of DNA damage in rainbow trout (Oncorhynchus mykiss) and gilthead sea bream (Sparus aurata) cryopreserved sperm

Cryopreservation causes several types of damage to spermatozoa, such as loss of plasma membrane integrity and functionality, loss of motility, and ATP content, resulting in decrease of fertility rates. This spermatozoal damage has been widely investigated for several marine and freshwater fish species. However, not much attention has been paid to the nuclear DNA. The objective of this study was to determine the degree to which cryopreservation induces spermatozoal DNA damage in two commercially cultured species, rainbow trout (Oncorhynchus mykiss) and gilthead sea bream (Sparus aurata), both of which could benefit from the development of cryopreservation strategies on a large scale. We have used the single-cell gel electrophoresis, commonly known as Comet assay to detect strand breaks in DNA. This technique was performed on fresh and cryopreserved sperm from both species. In rainbow trout there was a significant increase in the averages of fragmented DNA and Olive tail moment after cryopreservation (11.19-30.29% tail DNA and 13.4-53.48% Olive tail moment in fresh and cryopreserved sperm, respectively), as well as in the proportion of cells with a high percentage of DNA fragmentation. For gilthead sea bream there were no significant differences in the percentage of tail DNA between the control samples and sperm diluted 1:6 and cryopreserved (28.23 and 31.3% DNA(t), respectively). However, an increase in the sperm dilution rate produced an increase in the percentage of DNA fragmentation (41.4%). Our study demonstrates that cryopreservation can induce DNA damage in these species, and that this fact should be taken into account in the evaluation of freezing/thawing protocols, especially when sperm cryopreservation will be used for gene bank purposes.     

Synergistic effects of systemic trefoil factor family 1 (TFF1) peptide and epidermal growth factor in a rat model of colitis

Novel therapies for the treatment of colitis are required. We therefore examined the potential value of the trefoil factor family 1 (TFF1) peptide and epidermal growth factor (EGF) alone and in combination. Effects of TFF1- Cys58 +/- EGF on an in vitro HT29 cell wounding model of restitution showed synergistic activity when used in combination. In addition, animals had colitis induced by adding 4% dextran sulphate sodium (DSS) to the drinking water for 7 days and they also received twice daily subcutaneous injections of test peptides. Treatment with TFF1-Cys58 alone (100 microg/kg) reduced histological colitis score by 22%, but the TFF1-Ser58 variant was ineffective. In a second study, TFF1-Cys58 reduced histological colitis score by 15%, EGF (600 microg/kg) by 26%, and an additive response (42% reduction) was demonstrated when used together (P < 0.01 versus either peptide given alone). Similar results were found using tissue myeloperoxidase (MPO) activity as a marker of inflammation. Where clinical risk/benefit seems justified, these initial studies suggest that combination therapy of systemic EGF and TFF peptides may prove useful for treatment of colitis in patients with disease extending beyond the reach of topical (enema) therapy.     

Expression of a novel secretory form (Crb1s) of mouse Crumbs homologue Crb1 in skin development

Drosophila Crumbs and the mammalian homologues encoded by the Crb genes are transmembrane proteins required for determination of retinal cell polarity. We cloned a novel variant of mouse Crb1 and termed it Crb1s. Since the 3'-end of exon 6 remained unspliced, Crb1s coded for a short secretory protein lacking the transmembrane and cytoplasmic domains required for the function of Crb1. The Crb1 expression was confined to brain and eye, whereas Crb1s was detectable in various tissues including skin, lung, and kidney in adult mice. Active expression of Crb1s, but not Crb1, was observed during the skin development, in which localization of the Crb1s protein was altered from the basal layer to the upper layers. Cultured mouse keratinocytes synthesized the Crb1s protein and secreted a 80 kDa processed form to the supernatant. After Ca(2+)-induced differentiation, Crb1s became associated with focal adhesions and cell-cell contacts. Crb1s may play a role distinct from that of Crb1 in epidermal tissue morphogenesis.     

The use of flow cytometry in the evaluation of cell viability of cryopreserved sperm of the marine shrimp (Litopenaeus vannamei)

Although the cryopreservation of penaeid prawn sperm or embryos has definite applications in the aquaculture industry, there is no protocol routinely used for this procedure. One of the main problems relies on the limitations for the determination of sperm cell viability. In this study, we evaluated the toxicity and cryoprotectant effect of four agents, at three different concentrations, in sperm suspension, spermatic mass, and complete spermatophore of the marine shrimp Litopenaeus vannamei. Cells were frozen by fast and slow cooling rates. After thawing, they were analyzed by optical microscopy and flow cytometry, which was also utilized to determine spermatic viability by DNA staining with propidium iodine. Considering viability by morphotype analysis, the best result was obtained when the spermatic mass was frozen by slow cooling rate in the presence of methanol (61.6%). There was a positive correlation between morphotype analysis and flow cytometry, although the percentage of viable cells was always lower when determined by the later. These results show that flow cytometry is a valuable tool to evaluate sperm cell viability in decapod species and it is more sensitive technique than optical microscopy.     

Matrix mechanical properties of transversalis fascia in inguinal herniation as a model for tissue expansion

Inguinal herniation represents a common condition requiring surgical intervention. Despite being regarded as a connective tissue disorder of uncertain cause, research has focused predominantly on biochemical changes in the key tissue layer, the transversalis fascia (TF) with little direct analysis of functional tissue mechanics. Connective tissue tensile properties are dominated by collagen fibril density and architecture. This study has correlated mechanical properties of herniated TF (HTF) and non-herniated TF (NHTF) with fibrillar properties at the ultrastructural level by quasi-static tensile mechanical analysis and image analysis of collagen electron micrographs. No significant difference was found between any of the key mechanical properties (break stress, strain or modulus) for HTF and NHTF. In addition, no significant differences were found in average collagen fibril diameter, density or fibre bundle spacing. However, both groups displayed anisotropy with greater break stress (p=0.001) on average in the transverse anatomical plane compared to the longitudinal plane in a mean ratio of 2:1 (anisotropy ratio), though there was no evidence of a difference in this ratio for HTF and NHTF for both break stress and modulus. It was noted that this anisotropy ratio corresponds closely with the expected force distribution on a model cylindrical structure loaded axially. The absence of other functional differences does not support the idea of a failing (injured) tissue but is consistent with it being a tissue undergoing chronic growth/expansion under multi-vectored mechanical loading. These findings provide new clues to collagen tissue herniation for mathematical modelling and model tissue engineering.     

Metabolism Comparative Cytotoxicity Assay (MCCA) and Cytotoxic Metabolic Pathway Identification Assay (CMPIA) with cryopreserved human hepatocytes for the evaluation of metabolism-based cytotoxicity in vitro: proof-of-concept study with aflatoxin B1

Recently, we have improved the cryopreservation procedures for human hepatocytes, leading to cells that can be cultured after thawing (“plateable” cryopreserved human hepatocytes). The ability to culture cryopreserved human hepatocytes allows application of the cells for prolonged incubations such as long-term (days) metabolism studies, enzyme induction studies, and cytotoxicity studies. We report here the application of the plateable cryopreserved human hepatocytes to evaluate the relationship between xenobiotic metabolism and toxicity. Two assays were developed: The Metabolism Comparative Cytotoxicity Assay (MCCA) and the Cytotoxic Metabolic Pathway Identification Assay (CMPIA). The MCCA was designed for the initial identification of the role of metabolism in cytotoxicity by comparing the cytotoxic potential of a toxicant in a metabolically competent (primary human hepatocytes) and a metabolically incompetent (Chinese hamster ovary (CHO)) cell type, as well as the evaluation of the role of P450 metabolism by comparing the cytotoxicity of the toxicant in question in human hepatocytes in the presence and absence of a nonspecific, irreversible P450 inhibitor, 1-aminobenzotriazole (ABT). The CMPIA was designed for the identification of the P450 isoforms involved in metabolic activation via the evaluation of the cytotoxicity of the toxicant in the presence and absence of isoform-selective P450 inhibitors. Results of a proof-of-concept study with the MCCA and CMPIA with a known hepatotoxicant, aflatoxin B1 (AFB1), are reported. AFB1 is known to require P450 metabolism for its toxicity. In the MCCA, AFB1 was found to have significantly higher cytotoxicity in human hepatocytes than CHO cells, therefore confirming its requirement for biotransformation to be toxic. ABT was found to effectively attenuate AFB1 cytotoxicity, confirming that P450 metabolism was involved in its metabolic activation. In the CMPIA, AFB1 cytotoxicity was found to be attenuated by ketoconazole and diethyldithiocarbamate, but not by furafylline, quinidine, and sulfaphenazole. Results with the isoform-selective inhibitors suggest that the isoforms inhibited by ketoconazole (mainly CYP3A4) and diethyldithiocarbamate (mainly CYP2A6, and CYP2E1), but not the isoforms inhibited by furafylline (mainly CYP1A2), sulfaphenazole (mainly CYP2C9) and quinidine (mainly CYP2D6) are involved in the metabolic activation of AFB1. This proof-of-concept study suggests that MCCA and CMPIA with cryopreserved human hepatocytes are potentially useful for the evaluation of the relationship between human xenobiotic metabolism and toxicity.     

Tissue specific expression of antifreeze protein and growth hormone transgenes driven by the ocean pout (<Emphasis Type="Italic">Macrozoarces americanus</Emphasis>) antifreeze protein OP5a gene promoter in Atlantic salmon (<Emphasis Type="Italic">Salmo salar</Emphasis>)

Previous research aimed at producing genetically improved salmon broodstock for aquaculture led to the creation of two lines of transgenic Atlantic salmon using gene constructs that were derived in part from the ocean pout OP5a antifreeze protein (AFP) gene. One of the lines was produced using an OP5a AFP gene in which the 5′ region of the promoter was removed (termed t-OP5a-AFP), and the other line contains a growth hormone (GH) transgene (EO-1α) that consists of a chinook salmon GH cDNA driven by a truncated OP5a AFP promoter that is almost identical to that of the t-OP5a-AFP construct. The similarity of the promoter regions of these transgenes provided an opportunity to evaluate their tissue specific expression patterns. Expression of mRNA was evaluated using Northern blot and RT-PCR techniques. The results demonstrate that the AFP and GH trangenes were expressed in almost all body tissues, suggesting that the promoter region of the OP5a AFP gene lacks tissue specific elements. Northern analysis revealed that expression of the t-OP5a-AFP gene was considerably greater than that of the EO-1α GH transgene. Only the spleen tissue of the GH transgenics showed a visible band of hybridization. In contrast clear bands of hybridization were evident in all tissues, except for blood cells, of the AFP transgenics with heart, liver and brain tissue showing the highest levels of mRNA expression. This higher level of expression could be attributable to the presence of introns in the t-OP5a-AFP transgene. Since the GH transgenic salmon grow considerably faster than non-transgenics the low levels of GH transgene expression in this line were clearly sufficient to produce the desired rapid growth phenotype. In contrast the levels of AFP expression were inadequate to impart any improvement in the freeze resistance of the AFP transgenic salmon.     

Thermal stability,mechanical properties and reducible cross-links of rat tail tendon in experimental diabetes

Thermal stability (measured as isometric contraction force), biomechanical properties and reducible cross-links were measured in tail tendons from streptozotocin diabetic rats, with and without insulin treatment. After 10 days of diabetes the maximum thermal contraction force was unchanged, but the relaxation following the maximal contraction was retarded. After 30 days the maximum contraction force was increased and the relaxation rate was decreased. The maximum strength and stiffness of the tendons were increased after 10 days of diabetes and even more after 30 days. There was no change in the density of reducible cross-links. However, diabetes increased the amount of glucose attached to the lysine and hydroxylysine residues of collagen. Insulin treatment prevented all changes in thermal stability and mechanical properties. The results indicate that stabilization of collagen fibres in diabetes does not follow the same pattern as that seen in normal ageing.     

Effect of sericin supplementation on heat shock protein 70 (HSP70) expression,redox status and post thaw semen quality in goat

Cryopreservation results in substantial deterioration of heat shock protein 70 (HSP70) and ultra-structural changes in sperm organelles, resulting in a marked reduction in post-thaw semen quality. The present study was aimed to explicate the effect of sericin supplementation on expression profile of HSP70, redox status and post-thaw semen quality in Barbari goat. Five Barbari bucks were used to collect thirty semen ejaculates by using artificial vagina and each ejaculate was divided into three aliquots to which sericin was supplemented at 0% (Control), 0.25% (T1) and 0.50% (T2). Further, extended semen samples were equilibrated followed by their cryopreservation. Post-thaw semen characteristics, redox status of seminal plasma, enzyme leakage and HSP70 gene/protein expression in spermatozoa were assessed in all the groups. Per cent progressive motile spermatozoa, spermatozoa having intact plasma membrane (HOST + ve) and intact acrosomes in post-thaw spermatozoa were significantly (p < 0.01) higher in T1 and T2 as compared to control. A significant (p < 0.01) reduction in abnormal spermatozoa was found in T1 as compared to T2. Sericin supplementation significantly (p < 0.05) improved the antioxidative status (SOD, GST, CAT), reduced lipid peroxidation (MDA) and also prevented enzyme (ALT, LDH) leakage as compared to control samples. qRT-PCR results revealed that HSP70 mRNA expression was significantly (p < 0.01) upregulated in T1 and T2 group as compared to control. The positive effect of sericin on expression of HSP70 was further confirmed by immunoblotting followed by densitometry revealing higher expression in T1 and T2 compared to control. Inclusion of 0.25% w/v sericin in semen extender ameliorated the post-thaw semen quality by improving antioxidative status and minimizing the leakage of intracellular enzymes. Sericin supplementation had a beneficial effect on HSP70/HSP70 mRNA expression either by induction or by protection of HSP70/HSP70 mRNA as evident from the gene expression and immunoblotting studies.     

Antinociceptive effect of (S)-N-desmethyl trimebutine against a mechanical stimulus in a rat model of peripheral neuropathy

Trimebutine (2-dimethylamino-2-phenylbutyl 3,4,5-trimethoxybenzoate, hydrogen maleate) relieves abdominal pain in humans. In the present study, the antinociceptive action of systemic (S)-N-desmethyl trimebutine, a stereoisomer of N-monodesmethyl trimebutine, the main metabolite of trimebutine in humans, was studied in a rat model of neuropathic pain produced by chronic constriction injury to the sciatic nerve. Mechanical (vocalization threshold to hindpaw pressure) stimulus was used. Experiments were performed two weeks after surgery when the pain-related behaviour has fully developed. (S)-N-desmethyl trimebutine (1, 3, 10 mg/kg s.c.) produced dose-dependent antinociceptive effects on the nerve-injured and the contralateral hindpaw. The effect of the lowest dose (1 mg/kg s.c.) of (S)-N-desmethyl trimebutine on the nerve-injured paw was equal to that seen after a ten time stronger dose on the contralateral paw. The effect of (S)-N-desmethyl trimebutine (1 mg/kg) was not naloxone reversible. The results suggest that systemic (S)-N-desmethyl trimebutine may be useful in the treatment of some aspects of neuropathic pain.     

Expression of IGF-binding protein-1 phosphoisoforms in fasted rat skin and its role in regulation of collagen biosynthesis

Insulin-like growth factor-I (IGF-I) is an important stimulator of collagen and glycosaminoglycan (GAG) biosynthesis in tissues. IGF-I activity is modulated by a family of IGF-binding proteins (IGFBPs) with different IGF-I binding affinities. At least IGFBP-1 and IGFBP-2 are known as inhibitors of IGF functions. Some IGFBPs (IGFBP-1, IGFBP-3 and IGFBP-5) may undergo phosphorylation that dramatically increase their affinity for IGF. During fasting of animals there is a significant decrease of the collagen and GAG content of the skin, accompanied by a reduction of plasma IGF-I levels. However, in previous studies we showed that in the skin of fasted rats IGF-I as well as IGFBP-1 and IGFBP-2 expressions were not different, compared to control rat skin, although collagen content was significantly decreased. In the present study we show that fasted rat skin contains similar amounts of IGF-I, IGFBP-3 and IGFBP-1, although extract from fasted rat skin induced inhibition of collagen biosynthesis in cultured fibroblasts, compared to control rat skin extract. Western immunoblot analysis of control and fasted rat skin extracts, using anti-phosphoserine antibodies for immunoprecipitated IGFBP-1 and IGFBP-3, revealed that both proteins are present in phosphorylated form. Although no differences were found in the expression of phosphorylated IGFBP-3 between control and fasted rat skins, that of phosphorylated IGFBP-1 in fasted rat skin extract was higher than in control one. We suggest that there is an increased level of IGFBP-1 phosphoisoform in fasted rat skin, associated with increased affinity for IGF-I. The increase of phosphorylated IGFBP-1 in fasted rat skin tissue may augment IGF-I binding affinity for IGF and decrease its bioavailability for receptor interaction. This mechanism may prevent IGF-I dependent stimulation of fibroblasts to produce extracellular matrix components. The specific expression of IGFBPs and their phosphoisoforms in tissues may play an important role in regulation of IGF-I action during physiologic and pathologic responses.     

Glucagon-like peptide 1 (7-36) amide (GLP-1) and exendin-4 stimulate serotonin release in rat hypothalamus

Glucagon-like peptide 1 (7-36) amide (GLP-1) and exendin-4 are gastrointestinal hormones as well as neuropeptides involved in glucose homeostasis and feeding regulation, both peripherally and at the central nervous system (CNS), acting through the same GLP-1 receptor. Aminergic neurotransmitters play a role in the modulation of feeding in the hypothalamus and we have previously found that peripheral hormones and neuropeptides, which are known to modulate feeding in the central nervous system, are able to modify catecholamine and serotonin release in the hypothalamus. In the present paper we have evaluated the effects of GLP-1 and exendin-4 on dopamine, norepinephrine, and serotonin release from rat hypothalamic synaptosomes, in vitro. We found that glucagon-like peptide 1 (7-36) amide and exendin-4 did not modify either basal or depolarization-induced dopamine and norepinephrine release; on the other hand glucagon-like peptide 1 (7-36) amide and exendin-4 stimulated serotonin release, in a dose dependent manner. We can conclude that the central anorectic effects of GLP-1 agonists could be partially mediated by increased serotonin release in the hypothalamus, leaving the catecholamine release unaffected.     

Effects of mechanical loading on the expression of pleiotrophin and its receptor protein tyrosine phosphatase beta/zeta in a rat spinal deformity model

Mechanical loading of the spine is a major causative factor of degenerative changes and causes molecular and structural changes in the intervertebral disc (IVD) and the vertebrae end plate (EP). Pleiotrophin (PTN) is a growth factor with a putative role in bone remodeling through its receptor protein tyrosine phosphatase beta/zeta (RPTPβ/ζ). The present study investigates the effects of strain on PTN and RPTPβ/ζ protein expression in vivo. Tails of eight weeks old Sprague-Dawley rats were subjected to mechanical loading using a mini Ilizarov external apparatus. Rat tails untreated (control) or after 0 degrees of compression and 10°, 30° and 50° of angulation (groups 0, I, II and III respectively) were studied. PTN and RPTPβ/ζ expression were evaluated using immunohistochemistry and Western blot analysis. In the control group, PTN was mostly expressed by the EP hypertrophic chondrocytes. In groups 0 to II, PTN expression was increased in the chondrocytes of hypertrophic and proliferating zones, as well as in osteocytes and osteoblast-like cells of the ossification zone. In group III, only limited PTN expression was observed in osteocytes. RPTPβ/ζ expression was increased mainly in group 0, but also in group I, in all types of cells. Low intensity RPTPβ/ζ immunostaining was observed in groups II and III. Collectively, PTN and RPTPβ/ζ are expressed in spinal deformities caused by mechanical loading, and their expression depends on the type and severity of the applied strain.