Neuronal regulation of bone metabolism and anabolism: calcitonin gene-related peptide-, substance P-, and tyrosine hydroxylase-containing nerves and the bone

Bone alters its metabolic and anabolic activities in response to the variety of systemic and local factors such as hormones and growth factors. Classical observations describing abundance of the nerves fibers in bone also predict a paradigm that the nervous system influences bone metabolism and anabolism. Identification of the nerve-derived signaling molecules, capable of modulating cellular activities of the bone cells, facilitates a novel approach to study the biology of skeletal innervation. Many of the signaling molecules that may act as efferent agents on the bone cells fall into the category of neuropeptides. The present article reviews current understanding of the skeletal innervation and their proposed physiological effects on bone metabolism, with a special interest to calcitonin gene-related peptide (CGRP)-containing nerves fibers. CGRP is abundantly distributed in bone via sensory nerves, especially in the epiphyseal trabecular bones. Its in vitro actions to the cultured osteoblasts and osteoclasts, together with its in vivo localization, strongly support the paradigm that the nervous system influences bone metabolism. In addition, CGRP is recently shown to be expressed endogenously by the osteoblasts. Transgenic mice with osteoblasts overexpressing CGRP are characterized by increased bone formation rate and enhanced bone volume, suggesting that CGRP indeed acts on bone metabolism not only via nervous route but also via autocrine loop. The current article also reviews the distribution of nerve fibers containing substance P (SP), another sensory nerve-specific neuropeptide, and tyrosine hydroxylase (TH), the rate-limiting enzyme of catecholamine. The distinct effects of SP and catecholamines on the bone cells together with their in vivo influences manifested by experimental denervation studies suggest that the sensory and sympathetic nerves play important roles in bone metabolism.     

Functional interaction of G protein-coupled receptors of the adrenomedullin peptide family with accessory receptor-activity-modifying proteins (RAMP)

Adrenomedullin (AM), alpha- and beta-calcitonin gene-related peptide (CGRP), calcitonin (CT), and amylin are homologous polypeptides with overlapping biological actions such as vasodilatation and inhibition of bone resorption. They are brought about through receptors that include the CT receptor (CTR) and an initially orphan CT receptor-like receptor (CRLR) in association with receptor-activity-modifying proteins (RAMP)1, -2, and -3. Co-expression of CRLR with RAMP1 or -2 revealed CGRP or AM receptors, respectively. The CTR interacts with CT and does not require a known RAMP for functional expression. The same CTR is a CGRP/amylin or an amylin receptor upon co-expression with RAMP1 or -3, respectively. Interactions between CRLR and RAMP are thought to be required for their delivery to the cell surface. There, heterodimeric complexes between CRLR or CTR and the corresponding RAMP reveal high-affinity receptors for AM, CGRP, and amylin. Here we review the current knowledge on interactions of G protein-coupled receptors with defined associated proteins.     

Histological demonstration of increased vascular permeability in the dura mater of the rat

Neurogenic inflammation of the dura mater encephali has been suggested to play an important role in the pathophysiology of headaches. Although functional studies using extravasation techniques indicate an enhanced permeability of blood vessels after chemical or electrical stimulation of C-fibres supplying the dura mater, histological demonstration of leaky blood vessels is still a problem. We used the vascular labelling method combined with i.v. injection of colloidal silver solution to test the permeability increasing effect of intravenous administration of substance P, topical application of mustard oil or acidic phosphate buffer and local electrical stimulation of the exposed dura mater. Histological characteristics of increased vascular permeability were observed exclusively after mustard oil and acidic phosphate buffer. This observation may indicate different mechanisms of increased vascular permeability involving pinocytosis and formation of interendothelial gaps selectively visualized by the vascular labelling method.     

Nerve fibers innervating the cranial and spinal meninges: morphology of nerve fiber terminals and their structural integration

Pachymeninx and leptomeninx of cranial cavity and spine are considerably different in their collagenous fiber texture, cellular composition, vascularization, and innervation. The majority of meningeal nerve fibers terminate as free nerve endings whereas encapsulated and lamellated nerve terminals additionally occur in higher vertebrates including man. With respect to nerve fiber classification, arborization pattern, topography, and organization of the microenvironment at the termination site afferent and efferent nerve terminals are differentiated. Only the dura mater and the pial subcompartment of the leptomeninx possess the morphological prerequisites for neurogenic inflammation. In the current review, the results of morphological studies regarding the meningeal innervation including the sites of CSF (cerebrospinal fluid) production and absorption are discussed with emphasis on their structure-function relationships.     

Three-dimensional wall structure and the innervation of dental pulp blood vessels.

The involvement of neural components in plasma extravasation and blood flow in the dental pulp has been established by pharmacological and physiological studies. We review here the segmental constitution of pulp vessels and the possible involvement of neural components in both the contractility and permeability of the pulp vessels from a morphological viewpoint. Six vascular segments can be identified based on the morphology of peri-endothelial cells, such as smooth muscle cells and pericytes. These are: muscular arterioles, terminal arterioles, precapillary arterioles, capillaries, postcapillary venules, and collecting or muscular venules. The perivascular nerve forms a mesh with numerous terminal varicosities, some of which attach directly to arteriolar smooth muscle cells. This mesh can be seen by scanning electron microscopy, and indicates the important role of neural components in regulating the pulpal circulation. After administering norepinephrine (0.2 mg/kg/dog), the surface texture of the smooth muscle cells of pulp arterioles reveals marked irregularities, which are correlated with arteriolar contraction. The pericytes in larger postcapillary venules (diameter 20 microm or larger) also show irregularities, whereas no changes are seen in the pericytes of either smaller postcapillary venules or capillaries. The intercellular spaces of pericytes in the postcapillary venules are wide enough for leukocytes to pass through, and the occasional extravasation of leukocytes through venule walls can be seen under electron microscopy. The microvessels of healthy human dental pulp react weakly to selectins, indicating that apparently healthy dental pulp may be weakly inflamed. In rat dental pulp, CGRP-immunoreactive nerves and nerve terminals containing many granular vesicles supply the postcapillary venules more densely than the arterioles, which suggests the involvement of postcapillary venules in neurogenic inflammation in the dental pulp.     

偏头痛与脑血管发育异常、脑白质变性、脑腔梗关系分析

目的 :观察偏头痛患者脑血管情况及颅内病变情况。方法 :以住院偏头痛患者为研究对象、同一时期无偏头痛患者为对照,进行神经影像学比较。结果 :58例偏头痛患者中,有脑腔梗病灶19例(32.76%),有脑白质变性17例(29.31%),有脑血管异常37例(63.79%)。61例对照组中有脑腔梗病灶9例(14.75%),有脑白质变性5例(8.20%),有脑血管异常13例(21.31%)。2组比较差异均有统计学意义(P<0.05)。结论:偏头痛与脑白质病变、脑腔梗有关,血管因素可能参与偏头痛的发作。     

Distribution of CGRP in the minipig brainstem

For the first time, an in‐depth study has been made of the distribution of fibers and cell bodies containing calcitonin gene‐related peptide (CGRP) in the minipig brainstem using an indirect immunoperoxidase technique. The animals studied were not treated with colchicine. Cell bodies containing CGRP were found in 20 nuclei/regions of the brainstem. These perikarya were located in somatomotor, brachiomotor and raphae nuclei, nucleus ambiguus, substantia nigra, nucleus reticularis tegmenti pontis, nucleus prepositus hypoglossi, nuclei olivaris inferior and superior, nuclei pontis, formatio reticularis, nucleus dorsalis tegmenti of Gudden, and in the nucleus reticularis lateralis. Fourteen of the 20 brainstem nuclei showed a high density of immunoreactive cell bodies. In comparison with other species, the minipig, together with the rat, show the most widespread distribution of cell bodies containing CGRP in the mammalian brainstem. Immunoreactive fibers were also observed in the brainstem. However, in the minipig brainstem the density of these fibers is low, as in many brainstem nuclei only single immunoreactive fibers were observed. A high density of immunoreactive fibers was only observed in the pars caudalis of the nucleus tractus spinalis nervi trigemini and in the nucleus ventralis tegmenti of Gudden. According to the observed anatomical distribution of the immunoreactive structures containing CGRP, the peptide could be involved in motor, somatosensory, gustative, and autonomic mechanisms. Microsc. Res. Tech. 77:374–384, 2014. © 2014 Wiley Periodicals, Inc.     

Cholinergic-nitrergic transmitter mechanisms in the cerebral circulation

Cerebral blood vessels from several species are innervated by vasodilator nerves. Acetylcholine (ACh) released from parasympathetic cholinergic nerves was first suggested to be the transmitter for vasodilation. Results from pharmacological studies in isolated cerebral arterial ring preparations, however, have demonstrated that nitric oxide (NO) but not ACh mediates the major component of neurogenic vasodilation. More recently, ACh and NO have been shown to co-release from the same cholinergic-nitrergic nerves, and that ACh acts as a presynaptic transmitter in modulating NO release. In this communication, evidence for the neuronal origin of NO and possible role of ACh in modulating NO release in large cerebral arteries at the base of the brain will be discussed.     

鲑鱼降钙素合成中二硫键形成的反相高效液相色谱分析

多肽中二硫键是否存在,将有影响对该多肽在RP－HPLC的保留时间,本文介绍在鲑鱼降钙素的合成中二硫键形成前后,即氧化态与还原态在选定的RP－HPLC色谱条件下形成两个可分离的谱峰,利用两个谱峰峰面积作为氧化程度的一个评价指标。     

痛泻要方对肠易激综合征大鼠血浆及肠黏膜CGRP影响的实验研究

目的：探讨肠易激综合征（irritable bowel syndrome）的发病机制以及痛泻要方煎剂治疗肠易激综合征内脏高敏感性模型大鼠的机制。方法：选用清洁级新生sD大鼠,采用结肠慢性刺激法加夹尾刺激法造模。造模成功后,将实验动物分组为空白对照组（A组）、模型对照组（B组）、中药低剂量组（C组）、中药中剂量组（D组）、中药高剂量组（E组）、西药奥替溴铵（F组）。A组和B组以生理盐水按4ml／100g（40g／kg／d）灌胃,C、D、E、F组分别以低剂量痛泻要方煎剂组按0．4ml／100g（4g／kg／d）、中剂量痛泻要方煎剂组按1．2ml／100g（12g／kg／d）、高剂量痛泻要方煎剂组按4ml／100g（40g／kg／d）灌胃、西药奥替溴铵组按4ml／100g（150g,kg／d）灌胃,每天两次,连续30天。结果：痛泻要方高剂量组能显著增加血浆和结肠组织CCRP含量,痛泻要方低剂量组、中剂量组与西药组均不能。结论：痛泻要方可能是通过增加模型大鼠血浆和结肠组织CGRP含量来实现的。     

Molecular inflammation hypothesis of aging based on the anti-aging mechanism of calorie restriction

Accumulating evidence strongly suggests that oxidative stress underlies aging processes. Research provides consistent evidence that calorie restriction (CR) reduces age-related oxidative stress and has anti-inflammatory properties. However, information is lacking on the molecular mechanism that would better define the interrelation of reactive oxygen species and nitrogen species and the pro-inflammatory states of the aging process. In this review, the biochemical and molecular bases of the inflammatory process in the aging process are analyzed to delineate the molecular inflammation hypothesis of aging. The key players involved in the proposed hypothesis are the age-related upregulation of NF-kappa B, IL-1 beta, IL-6, TNFalpha, cyclooxygenase-2, and inducible NO synthase, all of which are attenuated by CR. Furthermore, age-related NF kappa B activation is associated with phosphorylation by I kappa B kinase/NIK and MAPKs, while CR blocked these activation processes. The modulation of these factors provides molecular insights of the anti-inflammatory action of CR in relation to the aging process. Based on available finding and our recent supporting evidence, we prefer to use "molecular inflammation" to emphasize the importance of the molecular reaction mechanisms and their aberrance, predisposing to fully expressed chronic inflammatory phenomena. It was further proposed that CR's major force of the regulation of redox-sensitive inflammation may well be its life-prolonging action.     

Quantification of structural changes in acute inflammation by fractal dimension,angular second moment and correlation

The aim of the study was to examine alteration and possible application of fractal dimension, angular second moment, and correlation for quantification of structural changes in acutely inflamed tissue. Acute inflammation was induced by injection of turpentine oil into the right and left hind limb muscles of mice, whereas control animals received intramuscular saline injection. After 12 h, animals were anesthetised and treated muscles collected. The tissue was stained by hematoxylin and eosin, digital micrographs produced, enabling determination of fractal dimension of the cells, angular second moment and correlation of studied tissue. Histopathological analysis showed presence of inflammatory infiltrate and tissue damage in inflammatory group, whereas tissue structure in control group was preserved, devoid of inflammatory infiltrate. Fractal dimension of the cells, angular second moment and correlation of treated tissue in inflammatory group decreased in comparison to the control group. In this study, we were first to observe and report that fractal dimension of the cells, angular second moment, and correlation were reduced in acutely inflamed tissue, indicating loss of overall complexity of the cells in the tissue, the tissue uniformity and structure regularity. Fractal dimension, angular second moment and correlation could be useful methods for quantification of structural changes in acute inflammation.     

Microvasculature in gingivitis and chronic periodontitis: disruption of vascular networks with protracted inflammation.

Gingivitis occurring when bacterial plaque accumulates in the gingival crevice provides a convenient and interesting model for chronic inflammation in humans. In some patients, gingivitis progresses to the destructive lesion of periodontitis, involving the formation of periodontal pockets. The basis for pocket formation and progression is not as yet clear, although neutrophilic polymorphonuclear leukocytes (PMN) appear to play a protective role. Vascular changes appear to either facilitate or inhibit PMN function with the effect of either protecting from, or stimulating, periodontitis. Contrary to most circumstances, high endothelial cells in periodontitis are involved with PMN rather than lymphocyte emigration. Expansion of the microvasculature through increased vascular diameter and tortuosity as well as the development of high endothelial cells appears to protect from periodontitis by increasing the supply of both plasma defense factors and PMN to the tissues. Vascular changes that may oppose this and promote periodontitis are the formation of perivascular hyaline material and accumulation of basement membrane rests. The inadequate tissue turnover that accumulation of these vascular products represents can be argued as a vascular response to a chronic inflammation that has failed to eliminate the irritant. It is suggested that these vascular changes may account for the highly localized and burst-like pattern of pocket formation in periodontitis. Finally, it is possible that the recent observation that periodontitis is an independent risk factor for systemic vascular disease may reflect stimulation of acute phase protein synthesis by cytokines released by periodontal high endothelial cells.     

Immunohistochemical characterization of TH13‐L2 spinal ganglia neurons in sheep (Ovis aries)

Spinal ganglia (SG) neurons are commonly classified according to various specific features. The most widespread classification based on morphological and ultrastructural features subdivides SG neurons into light and small dark neurons. Using immunohistochemical, histochemical and lectin methods, it is possible to further subdivide the small dark neurons into two subpopulations: peptidergic and nonpeptidergic neurons. The majority of studies on SG neurons were carried out on mice and rats; there are few or no studies on large mammals. In this study, some of the widely used neuronal markers, neurofilament 200 kDa (NF200), substance P (SP), calcitonin gene‐related peptide (CGRP) and isolectin B4 (IB4), were employed to characterize neuronal nitric oxide synthase (nNOS)‐immunoreactivity (‐IR) in sheep (Ovis aries) SG (Th13‐L2) neurons. The majority of the SG neurons were IB4‐labeled (79 ± 10%), followed by NF200‐ (45 ± 4%), NOS‐ (44 ± 10%), SP‐ (42 ± 5%) and CGRP‐IR (35 ± 7%) neurons. The triple staining experiments showed that a higher percentage (75 ± 16%) of NOS‐IR neurons bound both IB4 and CGRP, or both IB4 and SP (49 ± 6%). The IB4 marker showed an unexpected staining pattern; in fact, IB4‐labeled neurons largely colocalized with NF200, usually considered a marker of light SG neurons, and with CGRP and SP. For this reason, IB4 cannot be employed in sheep to differentiate between light and dark neurons, or between peptidergic and nonpeptidergic neurons. These results suggest the importance of being cautious when comparing data among different species. Microsc. Res. Tech., 2010. © 2009 Wiley‐Liss, Inc.     

Role of dendritic cells and Th2 lymphocytes in asthma: lessons from eosinophilic airway inflammation in the mouse

Asthma is a chronic disorder of the airways characterized by variable airway narrowing, mucus hypersecretion, and infiltration of the airway wall with eosinophils. It is now believed that asthma is controlled by Th2 lymphocytes producing cytokines such as IL-4, IL-5, IL-9, and IL-13. Animal models of eosinophilic airway inflammation and airway hyperreactivity have been developed to study the contribution of cells or mediators in the pathogenesis of asthma. In this review, we discuss the role of antigen presenting cells, CD4(+) and CD8(+) T lymphocytes, B lymphocytes, NK cells, and mast cells in the induction and maintenance of eosinophilic airway inflammation, mucus hypersecretion, and airway hyperreactivity.     

A histopathological study of the pulp of dogs’ teeth after induction of experimental pulp inflammation for different periods of time

The purpose of this study was to investigate the reaction of the pulp of dogs’ teeth after insertion of soft carious dentin from freshly extracted human teeth into the buccal cavities for short and longer periods of time. Forty‐seven mature lower and upper teeth were used in this study. On the middle of the buccal side of the teeth, 3 × 5 mm class ν cavities were prepared, soft carious dentin from freshly extracted human teeth was inserted into the floor of the cavity, and those were filled with glass ionomer. The dogs were killed after 7, 14, 28 and 47 days using vital perfusion techniques. Six‐micrometre sections were prepared and blindly evaluated by pathologist. An inflammatory reaction occurred in all of the samples. Moderate to severe inflammation were shown in all periods except in one in the 7‐day period. Insertion of soft carious dentin gathered from freshly extracted human teeth into class ν cavities produced in dogs’ teeth for 7 days is a reproducible process and could be used in study of pulp pathology.     

Asymptotic fractals in the context of grey-scale images

The estimation of the fractal dimension in the case of concave log–log Richardson–Mandelbrot plots can be obtained by using asymptotic fractal equations. We demonstrate here, under asymptotic fractal conditions, that additional derivations making use of the Minkowski dilation in grey-scales lead to two asymptotes, one having a slope of 1 and the other a slope of D_T ? D + 1 (where D_T is the topological dimension and D the fractal dimension). The resulting equation offers important advantages. It allows: (i) evaluation of scaling properties of a grey-scale image; (ii) estimation of D without any iteration and (iii) generation of texture and heterogeneity models. We concentrate here on the first two possibilities. Images from cultured cells in studies of cytoskeleton intermediate filaments and kinetic deformability of endothelial cells were used as examples.     

基于SolidWorks的曲轴动平衡分析

传统的曲轴零件设计是设计人员凭借自身经验进行设计,常导致其动平衡的计算精度不高.介绍在刚性转子动平衡计算分析方法的基础上,结合Delphi 7.0和SolidWorks环境进行的曲轴零件动平衡分析的二次开发.这种设计方法可以在曲轴的设计过程中直接对曲轴零件进行动平衡分析,提高计算精度并节省曲轴的设计开发时间和成本.