Histological and immunohistochemical analyses of articular cartilage during onset and progression of pre- and early-stage osteoarthritis in a rodent model.

Early diagnosis and treatment of pre- and early-stage osteoarthritis (OA) is important. However, the cellular and cartilaginous changes occurring during these stages remain unclear. We investigated the histological and immunohistochemical changes over time between pre- and early-stage OA in a rat model of traumatic injury. Thirty-six male rats were divided into two groups, control and OA groups, based on destabilization of the medial meniscus. Histological and immunohistochemical analyses of articular cartilage were performed on days 1, 3, 7, 10, and 14 postoperatively. Cell density of proteins associated with cartilage degradation increased from postoperative day one. On postoperative day three, histological changes, including chondrocyte death, reduced matrix staining, and superficial fibrillation, were observed. Simultaneously, a compensatory increase in matrix staining was observed. The Osteoarthritis Research Society International score increased from postoperative day seven, indicating thinner cartilage. On postoperative day 10, the positive cell density decreased, whereas histological changes progressed with fissuring and matrix loss. The proteoglycan 4-positive cell density increased on postoperative day seven. These findings will help establish an experimental model and clarify the mechanism of the onset and progression of pre- and early-stage traumatic OA.

Age-related changes in rat joint capsule.

[Purpose] We investigated morphological and histopathological changes in the joint capsule of rats with aging. [Materials and Methods] A total of 18 male Wistar rats were categorized into two groups: the control group (n=8), and the aged group (n=10). The aged group was reared until 75 weeks of age, while the control group was maintained until 11 weeks of age. At the end of the experiment period, the knee joints were sampled, joint capsules were subjected to histopathological analysis, and their thickness was measured. [Results] The joint capsule in the aging group exhibited significantly greater thickness compared to the control group. Histopathological examination revealed distinct differences between the two groups. The control group displayed gaps between the collagen fibers in the posterior joint capsule, along with loosely overlapping connective tissue and the presence of fat cells. Conversely, in the aged group's joint capsule, these gaps between the collagen fibers almost disappeared and fibers became densely packed and thickened. [Conclusion] These results were similar to our previous study in rats with immobilized hindlimb knee joints. Similar findings, including collagen fiber thickening, densification in the joint capsule, and reduced hindlimb knee joint range of motion, were consistent with those observed in the present investigation.

Treadmill Exercise Suppresses Histological Progression of Disuse Atrophy in Articular Cartilage in Rat Knee Joints during Hindlimb Suspension.

OBJECTIVE: The purpose of this study was to determine the preventive effects of treadmill exercise or physiological loading on disuse atrophy in the rat knee joint cartilage and bone during hindlimb suspension. DESIGN: Twenty male rats were divided into 4 experimental groups, including the control, hindlimb suspension, physiological loading, and treadmill walking groups. Histological changes in the articular cartilage and bone of the tibia were histomorphometrically and immunohistochemically evaluated 4 weeks after the intervention. RESULTS: Compared with the control group, the hindlimb suspension group showed thinning of cartilage thickness, decreased matrix staining, and decreased proportion of noncalcified layers. Cartilage thinning, decreased matrix staining, and decreased noncalcified layers were suppressed in the treadmill walking group. The physiological loading group exhibited no significant suppression of cartilage thinning or decreased noncalcified layers, but the decreased matrix staining was significantly suppressed. No significant prevention of bone mass loss or changes in subchondral bone thickness were detected after physiological loading or treadmill walking. CONCLUSION: Disuse atrophy of the articular cartilage caused by unloading conditions could be prevented by treadmill walking in rat knee joints.

Immunohistochemical study on the distribution of telocytes in the knee joint components in a rat osteoarthritis model.

[Purpose] Telocytes are stromal cells that participate in tissue homeostasis. Osteoarthritis is a common degenerative disorder of multiple joint components that causes inflammation; however, the distribution of telocytes in joint components and the impact of osteoarthritis on telocytes is unclear. Therefore, we aimed to clarify the distribution of the telocyte in the joint components and determine the effect of osteoarthritis on telocytes. [Participants and Methods] We divided 30 male rats into control and osteoarthritis groups and surgically induced osteoarthritis by destabilizing the medial meniscus. At two and eight weeks after surgery, we evaluated the changes in CD34-positive and CD31-negative area sizes in the joint components by immunohistochemistry. [Results] The results showed CD34-positive and CD31-negative areas in the loose connective tissue of the lateral meniscus attachment and the infrapatellar fat pad. However, it was not observed in the cartilage, subchondral bone, cruciate ligament, and meniscus. Moreover, there were no significant differences between the CD34-positive and CD31-negative area sizes in control and osteoarthritis groups at both time points. [Conclusion] CD34-positive and CD31-negative cells are distributed in multiple joint components; however, CD34-positive and CD31-negative areas are not affected by the progression of osteoarthritis. This result provides information on telocytes distribution in the knee joint and the impact of osteoarthritis on these cells.

Physiological Reloading Recovers Histologically Disuse Atrophy of the Articular Cartilage and Bone by Hindlimb Suspension in Rat Knee Joint.

OBJECTIVE: This study aimed to clarify physiological reloading on disuse atrophy of the articular cartilage and bone in the rat knee using the hindlimb suspension model. DESIGN: Thirty male rats were divided into 3 experimental groups: control group, hindlimb suspension group, and reloading after hindlimb suspension group. Histological changes in the articular cartilage and bone of the tibia were evaluated by histomorphometrical and immunohistochemical analyses at 2 and 4 weeks after reloading. RESULTS: The thinning and loss of matrix staining in the articular cartilage and the decrease in bone volume induced by hindlimb suspension recovered to the same level as the control group after 2 weeks of reloading. The proportion of the noncalcified and calcified layers of the articular cartilage and the thinning of subchondral bone recovered to the same level as the control group after 4 weeks of reloading. CONCLUSIONS: Disuse atrophy of the articular cartilage and bone induced by hindlimb suspension in the tibia of rats was improved by physiological reloading.

Reduction of knee joint load suppresses cartilage degeneration, osteophyte formation, and synovitis in early-stage osteoarthritis using a post-traumatic rat model.

The purpose of this study was to clarify the histological effect of reducing the loading to knee on cartilage degeneration, osteophyte formation, and synovitis in early-stage osteoarthritis (OA) using a post-traumatic rat model. Ten male rats were randomly allocated into two experimental groups: OA induction by surgical destabilization of medial meniscus (DMM, OA group) and hindlimb suspension after OA induction by DMM (OAHS group). The articular cartilage, osteophyte formation, and synovial membrane in the medial tibiofemoral joint were analyzed histologically and histomorphometrically at 2 and 4 weeks after surgery. The histological scores and changes in articular cartilage and osteophyte formation were significantly milder and slower in the OAHS group than in the OA group. At 2 and 4 weeks, there were no significant differences in cartilage thickness and matrix staining intensity between both the groups, but chondrocytes density was significantly lower in the OA group. Synovitis was milder in OAHS group than in OA group at 2 weeks. Reducing knee joint loading inhibited histological OA changes in articular cartilage, osteophyte formation, and synovial inflammation. This result supports the latest clinical guidelines for OA treatment. Further studies using biochemical and mechanical analyses are necessary to elucidate the mechanism underlying delayed OA progression caused by joint-load reduction.

Disuse Atrophy of Articular Cartilage Induced by Unloading Condition Accelerates Histological Progression of Osteoarthritis in a Post-traumatic Rat Model.

OBJECTIVE: The study aim was to evaluate the histological relationship between osteoarthritis (OA) and articular cartilage in disuse atrophy induced by hindlimb unloading in a post-traumatic OA rat model. DESIGN: Forty male rats were divided into the 4 following experimental groups: control, hindlimb suspension (HS), OA induced by destabilization of the medial meniscus (OA), and OA induction after hindlimb suspension (HS-OA). Histological changes in the articular cartilage of the tibia were evaluated by the Osteoarthritis Research Society International (OARSI) scores and histomorphometrical analyses at 2, 4, and 8 weeks after OA induction. RESULTS: We confirmed that disuse atrophy of the articular cartilage was caused by thinning of the articular cartilage and the decrease in matrix staining for the nonloading period of 4 weeks. The OARSI scores and histomorphological analyses revealed that OA progressed significantly wider and deeper in the HS-OA group than in the OA group over time. In the sham group, disuse atrophy of the articular cartilage recovered at 2 weeks after reloading. CONCLUSIONS: This study revealed that OA progressed faster in cartilage atrophy than in normal articular cartilage. Further studies are required for investigating the mechanisms of disuse atrophy of cartilage and its association with OA using the biochemical and immunohistochemical analysis.

Daily and short-term application of joint movement for the prevention of infrapatellar fat pad atrophy due to immobilization.

[Purpose] To mobilize the knee joint during cast fixation and to determine whether infrapatellar fat pad changes can be prevented. [Materials and Methods] We randomly allocated Wistar rats into 3 groups as follows: normal group, raised in normal conditions (n=5); contracture group, immobilized with cast fixation (n=5); and prevention group, treated with joint movement during immobilization (n=5). We immobilized the right hindlimb using cast fixation. Joint movement in the prevention group was accomplished by repeatedly pulling the right hindlimb caudally and then returning the leg to the bent position for 10 minutes every day for 2 weeks. We used a metronome to maintain a constant speed, with one set lasting 2 seconds (1-second traction and 1-second return). [Results] The contracture group had adipose cells of various sizes and fibrosis in the infrapatellar fat pad. These changes were also found in milder forms in the prevention group. We found significant differences in the cross section of adipose cells and in knee extension restriction between the groups. [Conclusion] Promoting joint movement may not only have a therapeutic effect on adipose cells but also a preventative effect.

Disuse histological changes of an unloading environment on joint components in rat knee joints.

Objectives: To clarify the effects of mechanical unloading on joint components, including the articular cartilage by knee compartments, synovial membrane, and the infrapatellar fat pad (IFP), using a hindlimb suspension rat model. Design: Twenty-five male rats were divided into the three following experimental groups: the baseline, control (CON), and hindlimb unloading (HU) groups. Rats in the HU group were subjected to periods of hindlimb unloading by tail suspension. The joint components were evaluated by histopathological and histomorphometric analyses. Results: The unloading condition caused articular cartilage thinning and decreased matrix staining. The tendency of these histological changes in articular cartilage was similar between the knee compartments. In general, the unloading environment resulted in no critical histological changes to the synovial membrane and IFP. At 4 weeks, thickening of the synovial membrane and synovial-like tissue invasion were observed in the HU group, but there was no significant change in inflammation. There was no obvious histological change to the IFP caused by unloading. Conclusion: The unloading environment caused disuse histological changes in the articular cartilage; however, the tendency of the changes was similar between compartments. Furthermore, the unloading condition caused no critical histological changes to the synovial membrane or IFP. Clinically, the unloading environment without joint immobilization over a short period may not have caused critical histological changes to the joint components.

Induction of osteoarthritis by injecting monosodium iodoacetate into the patellofemoral joint of an experimental rat model.

This study aimed to investigate the histopathological changes in the patellofemoral joint using a rat model of osteoarthritis that was induced using monosodium iodoacetate, and to establish a novel model of patellofemoral osteoarthritis in a rat model using histopathological analysis. Sixty male rats were used. Osteoarthritis was induced through a single intra-articular injection of monosodium iodoacetate in both knee joints. Animals were equally divided into two experimental groups based on the monosodium iodoacetate dose: 0.2 mg and 1.0 mg. Histopathological changes in the articular cartilage of the patellofemoral joint and the infrapatellar fat pad were examined at 3 days, 1 week, 2 weeks, 4 weeks, 8 weeks, and 12 weeks after the monosodium iodoacetate injection. In the 1.0-mg group, the representative histopathological findings of osteoarthritis were observed in the articular cartilage of the patellofemoral joint over time. Additionally, the Osteoarthritis Research Society International scores of the patellofemoral joint increased over time. The synovitis scores of the infrapatellar fat pad in both groups were highest at 3 days, and then the values decreased over time. The fibrosis score of the infrapatellar fat pad in the 1.0-mg group increased with time, whereas the fibrosis score in the 0.2-mg group remained low. Representative histopathological findings of osteoarthritis were observed in the articular cartilage of the patellofemoral joint in a rat model of osteoarthritis induced using monosodium iodoacetate. With appropriate selection, this model may be regarded as an ideal patellofemoral osteoarthritis model.

Effect of low-intensity pulsed ultrasound therapy on a rat knee joint contracture model.

[Purpose] Histopathological investigation of the effects of low-intensity pulsed ultrasound (LIPUS) on joint components using a rat knee joint contracture model. [Subjects and Methods] Nineteen, 9-week-old Wistar male rats were divided into a control group (n=6) and an experimental group. Rats in the experimental group underwent cast immobilization of the right rear limb for 8 weeks. They were then randomly divided into a non-treatment group (n=6), which was raised under normal conditions for 4 weeks, and a treatment group (n=7), which underwent LIPUS for 4 weeks. LIPUS irradiation was performed at a frequency of 3 MHz, an intensity of 30 mW/cm2, and a pulse rate of 20% duty cycle. Irradiation was performed once daily for 10 min, 5 days per week. At the end of this period, tissue specimens in which the knee sagittal plane could be observed were prepared and observed using an optical microscope. [Results] The extension-limiting angle of the knee joint was significantly less in the treatment group compared with the non-treatment group. The posterior joint capsule was significantly thicker only in the non-treatment group, and the density was 53.5 ± 7.5% for the control group, 77.2 ± 5.7% for the non-treatment group, and 69.2 ± 2.9% for the treatment group, with significant differences existing across all groups. [Conclusion] LIPUS may widen the space between collagen fiber bundles of the joint capsule, thereby improving the range of motion.

Long-term histopathological developments in knee-joint components in a rat model of osteoarthritis induced by monosodium iodoacetate.

[Purpose] This study was performed to evaluate the long-term histopathological changes in knee-joint components including synovial membrane and joint capsule in a rat model of osteoarthritis (OA) induced by monosodium iodoacetate (MIA). [Subjects and Methods] Fifty male rats were used. OA was induced through intra-articular injection of MIA, and ten rats were randomly allocated to each of five groups induced with OA for 1, 2, 4, 6, or 8 weeks. At the end of each period, the knee components were examined histopathologically. [Results] After 1 and 2 weeks, chondrocytes were weakly stained. After 4 weeks, fibrillation, fissuring, and eburnation were observed, whereas after 6 weeks, chondrocyte clustering and osteophyte formation were detected. In the synovial membrane, the proliferation of spindle-shaped cells and a multilayered structure of the surface cells were observed at 1 and 2 weeks, but the degree of these changes decreased over time. In the joint capsule, a narrowing of the space between collagen fiber bundles was observed at 4-8 weeks. [Conclusion] The long-term histopathological changes of the joint components observed in a rat model of OA induced by MIA were similar to those detected in OA, but differed at specific times and tissues.

An immunohistochemical study of the sciatic nerve in a rat knee immobilization model.

[Purpose] This study was performed to immunohistochemically evaluate changes in the periphery of the sciatic nerve in a rat model of knee immobilization, and to assess the effects of range of motion exercise. [Subjects and Methods] Twenty-one male rats were divided randomly into three groups: control (C), immobilized (I), and exercise (E group). Rats in the I and E groups had the right knee joint immobilized for 2 weeks. In the E group, range of motion exercise was also performed. After the experimental period, the periphery of the sciatic nerve was immunohistochemically observed. [Results] Immunohistochemical staining revealed that the myelin sheath and the perineurium in all groups were laminin positive. In the C and E groups, all rats showed normal staining. In contrast, 4 rats in the I group exhibited weak labeling. [Conclusion] Our results suggest that immobilization alters the perineurium at a molecular level and the range of motion exercise is essential for maintaining the environment of the perineurium.

Experimental joint immobilization and remobilization in the rats.

[Purpose] The purpose of our study was to clarify temporal effects on restrictions to range of motion and the histopathological changes of joint components after joint immobilization in a rat knee-contracture model. [Subjects] Fifty-four male Wistar rats were randomly divided into two groups: a fixation group, and a control group. [Methods] In the fixation group, unilateral knee joints were immobilized at full flexion using a plaster cast for 4 weeks. At four weeks the animals were randomly divided into six subgroups, corresponding to the time of examination after cast removal: 0, 4, 8, 16, 24, and 32 weeks. For comparison, control group animals of corresponding age were also examined. [Results] Although movement restrictions of the knee joint had completely recovered 6 weeks after the cast removal, cartilage and synovial membrane structures did not completely recover. [Conclusion] These findings have not previously been reported, and as they form an addition to the fundamental scientific foundations of physical therapy, further research must examine these findings from a variety of perspectives.

Differences in Cartilage Repair between Loading and Unloading Environments in the Rat Knee.

We investigated the histopathological and immunohistochemical effects of loading on cartilage repair in rat full-thickness articular cartilage defects. A total of 40 male 9-week-old Wistar rats were studied. Full-thickness articular cartilage defects were created over the capsule at the loading portion in the medial condyle of the femur. Twenty rats were randomly allocated into each of the 2 groups: a loading group and a unloading group. Twenty rats from these 2 groups were later randomly allocated to each of the 2 groups for evaluation at 1 and 2 weeks after surgery. At the end of each period, knee joints were examined histopathologically and immunohistochemically. In both groups at 1 and 2 weeks, the defects were filled with a mixture of granulation tissue and some remnants of hyaline cartilage. The repair tissue was not stained with toluidine blue in both groups. Strong staining of type I collagen was observed in the repair tissue of both groups. The area stained with type I collagen was smaller in the unloading group than in the loading groups, and the stained area was smaller at 2 weeks than at 1 week. In the staining for type II collagen, apparent staining of type II collagen was observed in the repair tissue of both groups at 1 week. At 2 weeks, there was a tendency toward a higher degree of apparent staining in the loading group than in the unloading group. Accordingly, these results indicated that loading and unloading in the early phase of cartilage repair have both merits and demerits.

Histopathological changes in the periphery of the sciatic nerve of rats after knee joint immobilization.

[Purpose] This study was performed to investigate the histological changes that occur in the periphery of the sciatic nerve in rats undergoing knee immobilization. [Subjects and Methods] 29 male 9-week-old Wistar rats were divided randomly into a control group (C group, n = 7) and an immobilized group (I group, n = 22). The animals in the I group had the left knee joint immobilized in maximal flexion with plaster casts for two weeks. After the experimental period, we obtained cross-sections of tissues from the center of the left thigh, and the periphery of the sciatic nerve was observed under an optical microscope after hematoxylin-eosin staining. [Results] In contrast to the rats of C group, the rats in I group showed adherence between the bundle of nerve fibers and perineurium, as well as thickening of the perineurium. These histological changes were statistically significant. [Conclusions] Immobilization of the knee joints of rats resulted in characteristic histological changes in the connective tissue around the sciatic nerve.

Influence of ROM Exercise on the Joint Components during Immobilization.

[Purpose] The aim of this study was to clarify the effects of the ROM exercise on joint components according to histopathological analysis. [Subjects and Methods] In total, twenty-six 9-week-old adult male Wistar rats were used in this study. The rats were randomly divided into three groups, the immobilization group (n=10), exercise group (n=10), and control group (n=6). The immobilization group and exercise group were anaesthetized and operated on under sterile conditions. The right knee joints in the immobilization group and exercise group were immobilized with external fixation at 120 degrees of flexion. Range of motion exercise was started from the day after immobilization. ROM exercise was performed in the exercise group once a day for 3 minutes, 6 days a week, for 2 weeks. [Result] The joint capsule in the immobilization group and exercise group showed narrowing of the collagen bundles in interstitial spaces but was less dense in the control group. In the immobilized group, a hyperplastic reaction was associated with infiltration into the articular cavity and adhesion to the surface of the articular cartilage. Conversely, in the exercise group, hyperplasia of tissue was localized to the synovial membrane. [Conclusion] This finding may suggest that ROM exercise induces some changes within the joint components and tissue metabolism.

A case of lymphocytic mastopathy requiring differential diagnosis from primary breast lymphoma.

Lymphocytic mastopathy is a benign breast disease characterized by dense fibrosis, lobular atrophy, and aggregates of lymphocytes in a periductal and perivascular distribution. The condition affects young to middle-aged women and frequently shows an association with diabetes mellitus or autoimmune disorders. Here, we report a case of the disease clinically and radiologically mimicking primary breast neoplasms. The patient was a 50-year-old woman without diabetes who presented with two firm lumps in her right breast. Breast imaging findings from mammography, ultrasonography, computed tomography (CT), and magnetic resonance imaging (MRI), respectively, revealed an abnormal appearance suspicious of malignancy. A core-needle biopsy specimen showed atypical accumulation of lymphoid cells, which was not easy to differentiate from primary breast lymphomas. Moreover, (18)fluorodeoxyglucose positron emission tomography (FDG-PET) examination detected abnormal uptake in the same lesions. Histological examination of a surgically obtained specimen showed characteristic appearance of lymphocytic mastopathy, which predominantly consisted of B-lymphocytes. In our case, it was difficult to distinguish this entity from breast cancer or low-grade B-cell lymphoma without surgical biopsy.

Effect of Immobilization on Insoluble Collagen Concentration and Type I and Type III Collagen Isoforms of Rat Soleus Muscle.

Immobilization is often associated with decreased muscle elasticity. This condition is known as muscle contracture; however, the mechanism remains unclear. The purpose of this study was to clarify the mechanism governing muscle contracture in rat soleus muscle by identifying changes in ankle joint mobility, insoluble collagen concentration and type I and type III collagen isoforms following 1- and 3-week immobilizations. Following a 1-week immobilization, range of motion (ROM) of dorsiflexion declined to 90% of the control value; additionally, ROM dropped to 67.5% of the control value after a 3-week immobilization. This finding suggested that ankle joint mobility decreases in conjunction with extended periods of immobilization. Insoluble collagen concentration in soleus muscles, which was unchanged after 1 week of immobilization, increased 3 weeks after immobilization. These results may be indicative of collagen fibers with strong intermolecular cross-links contained in the muscle was made increased relatively by 3 weeks of immobilization. Therefore, the change in intermolecular cross-links may be significant in terms of progress of muscle contracture with longer periods of immobilization. On the other hand, the ratio of type III to type I collagen isoforms in muscular tissue increased following a 1-week immobilization; moreover, this ratio remained constant after 3 weeks of immobilization. These data suggested that muscle immobilization may induce type III collagen isoform expression. The increase in the ratio of type III to type I collagen isoforms do not change in parallel with the increase in the limitation in ROM; however, this phenomenon probably is not closely related to the progress of muscle contracture. The change of collagen isoform in immobilized muscle may be involved in the mechanism governing the progression of muscle fibrosis.

[A case of adult neuroblastoma arising in the retroperitoneum following a poorly aggressive course].

A 69-year-old man complaining of enlarged cervical mass, appetite loss and lower abdominal pain was found to have abdominal tumors in heaps forming a large mass around the retroperitoneum. The biopsy specimen in the cervical mass showed undifferentiated carcinoma with neuroendocrine feature. This malignancy followed a poorly aggressive course and caused death in only 24 hospital days. The disease was diagnosed as undifferentiated neuroblastoma arising in the retroperitoneum by autopsy with appropriate immunohistochemical studies. Adult neuroblastoma in the peritoneum is rare and our case showed a aggressive behavior and unfortunate outcome.