膝关节骨关节炎滑膜中神经生长因子和P物质水平测试与观察

目的比较神经生长因子(nervegrowthfactor,NGF)和P物质(substanceP,SP)在膝骨性关节炎(osteoarthritis,OA)和单纯半月板撕裂膝关节滑膜中不同部位的表达。方法应用免疫组化的方法定性和半定量分析NGF和SP在膝OA(13例次)及单纯半月板撕裂患者(6例次)膝关节不同部位滑膜中的表达。用Gless神经纤维染色法观察滑膜内神经纤维的分布。结果NGF主要在滑膜的血管壁平滑肌层表达,OA膝内下方滑膜的NGF阳性血管率(67.2%)明显高于外侧滑膜(47.6%,P<0.05)和髌上囊处滑膜(29.9%)的表达以及单纯半月板撕裂膝关节内下方(38.9%)、外侧(41.0%)、髌上囊处(32.2%)滑膜的表达(P<0.01)。SP在滑膜血管壁上表达,OA膝内下方滑膜SP的表达(61.7%)明显高于外侧滑膜的表达(46.5%,P<0.05)和在髌上囊滑膜(31.6%)的表达以及单纯半月板撕裂膝关节内下方(43.1%)、外侧(40.5%)和髌上囊(33.2%)处滑膜的表达(P<0.01)。滑膜内的神经纤维分布广泛,在血管周围尤为明显,形成血管周围神经网。结论NGF和SP在膝OA滑膜内下方的表达明显高于它们在滑膜其它处的表达,与临床观察到的膝OA患者膝内下方疼痛和压痛部位一致。     

Distribution of neuropeptides in synovium of the knee with osteoarthritis

Synovial tissue was obtained from medial, lateral, and suprapatellar sites of 21 knees (15 patients) with medial compartmental osteoarthritis at surgery. All patients reported pain around the medial joint of their knees while walking and climbing stairs. For investigation of the synovial innervation, six samples were stained with modified gold chloride and the others with an immunohistochemical method using antisera against neuropeptides. The extent of synovitis in each part was scored using a new 10-point scale. The results showed that the synovium had an extensive neural network in the somatic and autonomic nervous systems. Neuropeptides were most abundant, with an especially large number of substance P and calcitonin gene related peptide immunoreactive free nerve endings. Some of the substance P positive nerve endings were surrounded by monocytes. Substance P and calcitonin gene related peptide were found more frequently in the medial than in the lateral or suprapatellar areas. Substance P positive free nerve endings showed more dendritic morphologic features in the medial region than did those in the lateral and suprapatellar regions, and small nerves were accompanied by newly developed vessels in synovial villi. In the medial region, the synovitis was more remarkable than in the lateral region. These findings suggest that free nerve endings containing substance P may modulate inflammation and the pain pathway in osteoarthritis.     

Increased cellular infiltrate in inflammatory synovia of osteoarthritic knees

OBJECTIVE: To determine the nature and origin of synovial inflammation in knees with osteoarthritis (OA). METHOD: Synovial samples were obtained from 21 medial compartmental knee OAs from 19 patients. First, using 11 medial knee synovial samples from 9 patients, a quantitative estimation of synovitis was made with both ordinary and immunohistochemical staining. Second, from the other 10 knees, tissue samples were taken from both the medial and the lateral compartments to quantify cells that infiltrated into the synovium. Medial synovium was immunostained using antibodies to anti-type II collagen, CD68, CD2, CD4, CD8, CD15, CD19, CD25, HLA-DR, CD1a and LN5. The lateral synovium was immunostained with anti-type II collagen, CD68, HLA-DR and CD4 antibody as a control. RESULT: Denatured cartilaginous detritus was found captured by synovial lining cells with a strong immunoreactivity to CD68 antibody, and whose phagocytic potential was activated. The number of anti-type II collagen-positive fragments in the medial compartment of the knee was larger than that found in the lateral compartment. Moreover, the population of CD68-positive cells in synovial tissue and HLA-DR-positive cells in the lining layer was larger in the medial compartment than in the lateral compartment. The number of CD4-positive cells (defined as helper/inducer T lymphocytes) was greater in medial synovium than in lateral synovium. CONCLUSION: Overall, this study strongly supports the concept that the synovitis observed in patients with knee OA might be induced by an immunological mechanism involving, to some extent, a macrophage/helper T cell interaction.     

Angiogenesis in two animal models of osteoarthritis

OBJECTIVE: We have previously described angiogenesis at the osteochondral junction and in synovium of knees from patients with osteoarthritis (OA), but little is known about how closely animal models of OA resemble human disease with respect to vascular growth. This study aimed to characterise two animal models of knee OA with particular respect to osteochondral and synovial angiogenesis. METHOD: We examined the spontaneous Dunkin-Hartley (DH) guinea pig and medial meniscal transection (MNX) rat models of OA. Vessels at the osteochondral junction and in the synovium were identified by lectin immunohistochemistry and quantified by computer-assisted image analysis. Disease severity was assessed using a scoring system. RESULTS: Blood vessels crossed the osteochondral junction in juvenile rats and guinea pigs, with higher densities in the lateral than medial tibial plateau, the number decreasing with maturation in the absence of other OA changes. In the rat model, increased vascular density was observed both at the osteochondral junction and in the synovium, whilst osteochondral vascularity in control rats decreased with maturation, OA rats showed a persistence of blood vessels at the osteochondral junction. In rat synovium, blood vessel fractional area was increased in the hypertrophied synovium 14 days after surgery, then decreased to control levels by day 28. Significant differences in vascularity were not observed between affected (medial) and spared (lateral) compartments of guinea pig knees. CONCLUSION: The rat meniscal transection model of OA reproducibly displays both osteochondral and synovial angiogenesis comparable to our previous observations in human knee OA. DH guinea pigs, by contrast, display low vascularity throughout their protracted course of OA development. Changes in vascularisation occur early during the development of OA in the rat, and may contribute to the pathogenesis of OA.     

Ultrastructure of sensory nerve endings in monkey (Macaca fascicularis) knee joint capsule

Ultrastructural studies of sensory endings in monkey posterior medial knee joint capsule were undertaken. Three distinct sensory nerve endings have been identified: free nerve endings, Ruffini corpuscles, and Pacinian corpuscles. The free nerve endings are present in all layers of the joint capsule excluding the synovium. Two types of Ruffini corpuscles have been found in the fibrous layer. The first type is characterized by a thin perineurial capsule, the second type by a thicker perineurial capsule, and extensive intracapsular space. Both types of Ruffini corpuscles are innervated by approximately one to four myelinated axons which lose their sheaths as they course through the corpuscle. They terminate on collagen fiber bundles as distinct swellings with spiny membrane projections that are covered by a thin basal lamina. These terminals contain abundant mitochondria, agranular vesicles, and irregularly arranged neurofilaments and neurotubules. Two types of Pacinian corpuscles were occasionally observed. The first was a small, typically laminated structure with an inner core at the layer between the synovium and the fibrous layer and between the fibrous layer and muscle/ligament; larger Vater-Pacinian corpuscles were noted only at the boundary between the fibrous layer and the muscle/ligament layer.     

Substance P immunoreactive nerve fibers of the canine laryngeal mucosa

Substance P (SP) immunoreactive nerve endings in the laryngeal mucosa were studied by PAP immunohistochemistry with light and electron microscopy. SP immunoreactive sensory endings were observed in the epithelium as intra-epithelial free nerve endings and taste bud-like structures. A small number of autonomic SP immunoreactive nerve fibers were observed running parallel to arterioles which were over 30 micron in diameter and terminated in glandular cells. Contrary to findings by silver impregnation, intraepithelial free nerve endings were more frequently observed on the lower surface of the vocal cord. The taste bud-like structures were classified into two different types: simple terminations and reticular terminations, according to the mode of the SP immunoreactive nerve fiber. Immature or degenerated taste bud-like structures in the larynx were assumed to be mechanical receptors because these receptors lacked outer taste pores and taste hairs.     

Neurohistology of the sinus tarsi and sinus tarsi syndrome

The purpose of this study was to clarify the neural anatomy of the sinus tarsi. The nerve endings of the synovium in the sinus tarsi were examined. The synovial membrane in the sinus tarsi was excised in 20 patients with sinus tarsi syndrome (20 feet) and in 2 feet from subjects without symptoms (controls). In 15 of the 20 patients and the two controls, the excised synovial membrane was studied histologically with staining by a modified Gairns gold chloride method. Numerous neural elements were observed in the sinus tarsi in all examined synovium. There were abundant free nerve endings and three types of mechanoreceptors: Pacinian corpuscles, Golgi corpuscles, and Ruffini corpuscles. Macroscopic observation and histological examination, using hematoxylin and eosin, in the other 5 patients revealed chronic synovitis in the sinus tarsi. Our findings suggest that the sinus tarsi is not only a talocalcaneal joint space but a source of nociceptive and proprioceptive information on the movement of the foot and ankle. Sinus tarsi syndrome may result from disorders of nociception and proprioception in the foot. Received for publication on Dec. 14, 1998; accepted on Feb. 23, 1999     

Hyaluronan suppressed nitric oxide production in the meniscus and synovium of rabbit osteoarthritis model.

Nitric oxide (NO) plays an important role in cartilage degeneration, and NO donors induce meniscus degeneration and synovium inflammation. This study evaluated the effect of intraarticular injections of hyaluronan (HA) on NO production in meniscus and synovium using an experimental osteoarthritis (OA) model. Thirty-six New Zealand white rabbits underwent unilateral anterior cruciate ligament transection (ACLT), and were divided into three groups. Four weeks after ACLT, the HA group started to receive intraarticular HA injections once a week for 5 weeks; the vehicle group started to receive the carrier of HA; and the no injection group, no treatment. All ACLT knees were harvested at the 9th week. Meniscus and synovium sections were examined by immunohistochemistry for nitrotyrosine. The pieces of these two tissues were cultured for 24 h. Culture supernatants were analyzed for nitrite concentration. The amount of NO produced by the meniscus was much larger than that produced by the synovium. NO productions in the meniscus and synovium of the HA group were significantly lower than those of the other groups. The results suggest that the inhibition of NO production in meniscus and synovium might be a part of the mechanism of the therapeutic effect of HA on OA.     

Nociceptive sensory innervation of the posterior cruciate ligament in osteoarthritic knees

Introduction

Although the posterior cruciate ligament (PCL) is considered to contain not only proprioceptive but also nociceptive sensory fibers, there is a lack of information about nociceptive sensory innervation of the PCL. We hypothesized that the PCL has constant nociceptive sensory innervation, suggesting the possible source of osteoarthritic (OA) knee pain.

Materials and methods

Innervation of the PCL was examined by immunohistochemistry with particular reference to nociceptive nerve fibers in OA knees. Sensory nerve fibers were semi-quantitatively counted in the PCL of OA knees, comparing with non-OA knees. Protein gene product 9.5 (PGP9.5) as a general neuronal marker and calcitonin gene related peptide (CGRP) as a marker for nociceptive neuron were used.

Results

The PCLs had constant CGRP-immunoreactive (IR) nerve fibers in both OA and non-OA knees. The difference of the CGRP-IR nerve density between groups did not reach a statistical significance (p = 0.062). For PGP9.5-IR nerve fibers, however, the PCLs in OA knees were statistically less innervated than non-OA knees (p = 0.0009).

Conclusions

Our results showed that, in spite of a significant decrease in total innervation in OA knees, the PCLs have constant nociceptive sensory innervation. Although the relationship between the decrease in total innervations in the PCL and OA pathophysiology is still unclear, the PCL is the possible source of OA knee pain. Our results should be taken into account when examining the pain source of the OA knees and handling the PCL during total knee arthroplasty.     

Innervation of nociceptors in the menisci of the knee joint: an immunohistochemical study

Using histology, we studied the innervation of nociceptors in the medial and lateral menisci of the knee joint. Specimens examined were taken from 16 patients during arthroplasty. The patients were 6 men and 10 women, with ages ranging from 14 to 76 years (mean 56 years). Immunohistochemistry with the unlabeled antibody biotin-streptavidin method was employed to detect protein gene product 9.5 (PGP 9.5) or substance P (SP) in the specimen. The antibody for PGP 9.5 detected nerve tissues in the menisci. Most but not all of the nerve fibers were associated with blood vessels. Nerve fibers and sensory receptors were found mainly in the peripheral, vascular zone, representing the outer one-third of the meniscus, and the innervated area was wider in the anterior and posterior horns. Pacinian and Ruffini corpuscles as well as free nerve endings were identified in these areas. Larger fibers coursed circumferentially in the peripheral zone, with smaller branches of nerve fibers running radially into the meniscus. Nerve fibers positive for SP were also detected in the menisci, but were fewer in number. Their branches also were fewer, oriented radially and paralleling blood vessels. This study showed that some of the pain in cases of meniscal tear could originate in the meniscus itself, especially with peripheral tears that may be accompanied by bleeding. Received: 2 June 1998     

Arthroscopic management of synovial chondromatosis of the knee. Findings and results in 18 cases

We report the results of arthroscopic removal of loose bodies and abnormal synovium from 18 knees with primary synovial chondromatosis. After a mean of three years, six months (range one to 10 years), 14 knees were either symptom-free or had only minor symptoms. Three of these had required two arthroscopic operations. Three patients were improved but not cured and there was one failure. The results were better than the published results of open operation for this condition. Three patterns of macroscopic appearances were noted: four knees had large lesions covered by normal synovium, 10 had small fragments of cartilage lying in or on the synovium and four had only free fragments of cartilage in the joint cavity but none in, on, or under, the synovium. These three appearances may represent three different disease processes.     

Correlations between both the expression levels of inflammatory mediators and growth factor in medial perimeniscal synovial tissue and the severity of medial knee osteoarthritis

An enhanced expression of the inflammatory mediators in the perimeniscal synovium in knee osteoarthritis (OA) has been suggested to contribute to progressive cartilage degeneration. However, whether the expression levels of these molecules correlated with the severity of OA still remained unclear. Medial perimeniscal synovial samples were obtained from 23 patients with Kellgren-Lawrence (K/L) grades 2 to 4 of medial knee OA. Immunohistochemical analysis of the synovium revealed that the MMP-1, COX-2 and IL-1β expression of the patients with K/L 4 to be significantly reduced in comparison to those with either K/L 2 or 3, while the TGF-β expression showed the opposite. The synovial expression of MMP-1 and IL-1β showed a significant negative correlation with the severity of OA, while that of TGF-β again showed the opposite. In conclusion, although synovial inflammation remained active, the MMP-1, COX-2 and IL-1β expression in synovium decreased depending upon the severity of OA, while the TGF-β expression increased.     

Assessment of the efficacy of joint lavage in rabbits with osteoarthritis of the Knee

We investigated the efficacy of joint lavage to alter the progression of the disease process in a rabbit model of knee OA in varying stages of the disease. Thirty‐three white rabbits were operated to induce OA, and then the rabbits were divided into three groups (10 in each) randomly. In group 1, on week 1 after the operation, joint lavage was performed on both knees in five rabbits as treatment group, and the other five rabbits were used as control group received no intervention. By the end of week 2 after operation, the rabbits were sacrificed. In group 2, five received joint lavage on week 2, and all were sacrificed on week 3. In group3, five received joint lavage on week 3, and all were sacrificed on week 4. In each group, histological evaluation showed that both the breakdown of articular cartilage and the inflammation of synovium were less in the knees treated with joint lavage than that in the control knees. The enzyme‐linked immunosorbent assay revealed that the expression of IL‐1β and TNF‐α in synovial fluid decreased significantly in the treatment group. Our findings suggested that joint lavage was beneficial for OA at different phases of OA in rabbit models. Joint lavage may be a beneficial method for the treatment of OA clinically. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:91–96, 2009     

Dietary vitamins and selenium diminish the development of mechanically induced osteoarthritis and increase the expression of antioxidative enzymes in the knee joint of STR/1N mice

OBJECTIVE: To study the influence of dietary vitamins and selenium on mechanically-induced osteoarthritis (OA) and the expression of antioxidative enzymes in male STR/1N and Balb/c mice. Male STR/1N mice are prone to develop OA caused by a varus deformity-induced mechanical overload of the medial tibial plateau. METHODS: After 12 months of feeding (special diet supplemented with the vitamins E, C, A, B6, B2, and selenium) serial histological sections of the knee joints were evaluated for development of osteoarthritic changes (grade 0-4). Serum glutathione peroxidase activity (GSH-px) was measured photometrically. Expression of antioxidative enzymes was demonstrated by immunohistochemistry. RESULTS: All control STR/1N mice showed OA lesions (grade 3-4) while the special diet decreased OA incidence significantly down to approximately 65% (mostly grade 2). Even in Balb/c mice the incidence was decreased by the special diet from approximately 21% (control animals; grade 1) to approximately 14%. Serum GSH-px activity increased diet-dependently in both mouse strains but was generally higher in Balb/c mice. In both mouse strains the special diet increased the expression of GSH-px and Cu/Zn-SOD in articular cartilage while there was no expression of Mn-SOD. There was also a special diet-dependent increase in expression of GSH-px in the synovium of both mouse strains while an increase in expression of Mn-SOD and Cu/Zn-SOD could only be seen in the synovium of STR/1N mice. CONCLUSIONS: A diet supplemented with vitamins/selenium might be important in prevention or therapy of mechanically induced OA. We hypothesize that free oxygen radical species might be involved in the mechanical induction of OA.     

The ratio of medial to lateral tibial plateau bone mineral density and compartment-specific tibiofemoral osteoarthritis

INTRODUCTION: Bone is thought to play an important role in osteoarthritis (OA) pathophysiology. Our aim was to look at specific features of OA and their relation to the ratio of medial:lateral tibial plateau bone mineral density (M:L BMD Ratio). METHODS: We examined our research question in the Framingham OA Study Cohort. All participants had BMDs and weight-bearing plain radiographs of the knees (2002-2005). M:L BMD Ratios were calculated using BMD from medial and lateral regions in the tibial plateau. Knee x-rays were read for osteophytes (OSTs), joint space narrowing (JSN), and sclerosis (Osteoarthritis Research Society International (OARSI) scoring system). Knees were classified as having medial and/or lateral JSN if they had JSN >or=1 in the medial and/or lateral tibiofemoral compartments, respectively. Medial and/or lateral OSTs were defined as medial and/or lateral tibial and/or femoral OSTs >or=2, respectively. Medial sclerosis and lateral sclerosis were defined as medial and lateral tibial sclerosis >or=1, respectively. We performed a logistic regression with medial JSN as the outcome and with M:L BMD Ratio groups as predictor variables, using the median group as the referent. Analyses were adjusted for age, sex, and body mass index (BMI). Generalized estimating equations were used to adjust for correlation between knees. Identical analyses were performed with medial OSTs, medial sclerosis, lateral JSN, lateral OSTs, and lateral sclerosis as the outcomes. RESULTS: Mean age of 1612 subjects (3048 knees) was 63.9 (Standard Deviation (SD)+/-8.9), 56% were women, and mean BMI was 28.5 (SD+/-5.5). M:L BMD Ratio was positively associated with medial JSN (P for linear trend <0.0001) and negatively associated with lateral JSN (P for linear trend <0.0001). The relations of the ratio with medial and lateral OSTs were j-shaped with P for quadratic trends <0.0001. There were also strong associations between M:L BMD Ratio and compartment-specific sclerosis (P for linear trends <0.0001) with most knees with medial and lateral sclerosis being in the highest and lowest M:L BMD Ratio groups, respectively. CONCLUSION: In summary, the extremes of the M:L BMD Ratio are strongly associated with individual radiographic features of OA. These findings add to existing evidence supporting the importance of understanding bone in OA pathophysiology.     

腰椎后关节滑膜皱折韧带的神经末梢的病理观察

目的 观察腰椎后关节部位的痛觉感觉器-游离神经末梢,方法 对10例腰椎间盘突出伴有腰椎后关节紊乱患者,在其椎间盘手术中留取椎后关节及其滑膜标本,分别采用HE染色法、硝酸银染色法、氯化金染色法、免疫组化法作病理观察。结果 滑膜皱折、韧带部均具有游离神经末梢。结论 解释了腰椎后关节紊乱症的疼痛位、疼痛性质在患者主观描述中模糊不清的原因。     

Early changes in lapine menisci during osteoarthritis development: Part I: cellular and matrix alterations

OBJECTIVE: Osteoarthritis (OA) is the most common form of arthritis and patients with meniscal and ligament injuries of the knee are at high risk to develop the disease. The purpose of this study was to evaluate changes occurring in both medial and lateral menisci from the knees of anterior cruciate ligament (ACL) transected rabbits during the early stages of OA development. DESIGN: Meniscal tissues from control and experimental rabbits were processed for histology and immunohistochemistry for assessment of matrix organization and composition. RESULTS: At 3 and 8 weeks following ACL transection, histological examination demonstrated extensive extracellular matrix deterioration. Altered cell distribution, areas depleted of cells, and areas of cell clusters were found within the medial but not in the lateral meniscus. Immunohistochemistry of both medial and lateral menisci demonstrated significant changes in collagen distribution. Type I and III collagen staining was increased in both medial and lateral menisci. In contrast, type II collagen staining was overtly increased only in the medial meniscus. CONCLUSION: These results demonstrate that after ACL transection, extracellular matrix deposition as well as altered matrix organization and altered cell distribution occur early in the medial meniscus.     

Clinical value of (99m)Tc-methylene diphosphonate (MDP) bone single photon emission computed tomography (SPECT) in patients with knee osteoarthritis

OBJECTIVE: To evaluate that single photon emission computed tomography (SPECT) could reflect the clinical severity of knee osteoarthritis (OA) and to determine the clinical usefulness of SPECT as a sensitive imaging method detecting early OA. METHODS: Sixty knees of thirty OA patients were examined with (99m)Tc-methylene diphosphonate (MDP) bone SPECT, the intensity of uptake was graded and the sites of uptake were localized. One hundred millimeter visual analog scale (VAS), Western Ontario McMaster Universities (WOMAC) Osteoarthritis Index scores and physical examination were assessed. Simple radiograph and musculoskeletal ultrasonography (US) were performed, and Kellgren-Lawrence (K-L) grades were determined. The relationships between clinical findings, simple radiographic findings, US findings and the intensity of uptake in SPECT were analyzed. RESULTS: There were increased uptakes in 91.7% of the knees, and patella and medial tibial condyle were the most common sites of the uptakes. Symptomatic knees had significantly higher intensity of uptake than asymptomatic knees. Uptake intensity of medial compartment in SPECT well correlated with clinical findings, such as VAS, WOMAC scores, and physical examination, as well as US findings. Even in the symptomatic knees without abnormal radiographic findings, the increased uptake was observed in SPECT. CONCLUSION: In patients with knee OA, SPECT findings are well correlated with clinical findings, such as pain scores and physical examinations, and SPECT appears to be a sensitive tool for early detection of knee OA. SPECT information could be useful for determining clinical severity of knee OA and for diagnosing early OA more effectively.