Changes in type I, II, and X collagen immunoreactivity of the mandibular condylar cartilage in a naturally aging rat model

The effect of aging on type I, II, and X collagen in the mandibular condyle was histologically and immunohistochemically assessed in 1−, 4−, 9−, and 16-month-old rats. Hypertrophic chondrocytes, observed in the 4-month-old rat, were absent in the 9-month-old. In 9- and 16-month-old rats, a mineralizing front ran parallel to the surface of the condyle, and the calcified cartilage was thicker than in the younger rats. Type I collagen was observed from the fibrous layer to the upper maturative cell layer in the 1- and 4-month-old rats. In the 9-month-old, the type I collagen-positive area extended to the whole cartilaginous region. In the 16-month-old, type I collagen took on an archlike configuration around the lacunae. Intense type II collagen reactivity in the maturative and hypertrophic cell layers of the 1-month-old rat was only slightly changed in the 4-month-old. In the 9-month-old rat, immunoreaction was detected from the proliferative cell layer; this extended to the whole cartilage in the 16-month-old. Type X collagen was localized in the hypertrophic cell layer in the 1-month-old and had expanded over the maturative cell layer in the 4-month-old rat. It was detected beneath the proliferative cell layer in the 16-month-old. Type X collagen was always observed in the area immediately above the mineralizing front of the cartilage matrix. Thus, our study indicated that mandibular condylar cartilage becomes fibrocartilage-like tissue with advancing age and that type X collagen may play a pivotal role in the progression of the mineralized front.     

Expression of collagen type I, II and III in loose body of osteoarthritis

The expression of collagen type I, II, and III was investigated to evaluate phenotypic change in chondrocytes in loose bodies related to osteoarthritis. We assessed collagen type I, II, and III production in loose bodies from knee joints of ten osteoarthritic patients, using an immunohistochemical method with monoclonal antibodies. Collagen type III expression was identified in all ten loose bodies and was mainly located in cartilage, including chondrocytes and matrices, as well as in a layer of fibroid tissue on the surface. No positive signal for collagen type III was observed in necrotic osteocytes. There was weakly positive staining for collagen type I in chondrocytes. No positive staining for collagen type II could be seen in the cartilage of loose bodies. Cartilage from the non-osteoarthritic knee joints of four people was negative for the expression of collagen type I and III, and positive for the expression of collagen type II. Collagen type I and III expression suggested the dedifferentiation status of chondrocytes in loose bodies. Received for publication on May 26, 1999; accepted on Oct. 20, 1999     

Low‐intensity pulsed ultrasound (LIPUS) increases the articular cartilage type II collagen in a rat osteoarthritis model

In this study, the effect of low‐intensity pulsed ultrasound (LIPUS) on cartilage was evaluated in a rat osteoarthritis (OA) model using serum biomarkers such as CTX‐II (type II collagen degradation) and CPII (type II collagen synthesis) as well as histological criteria (Mankin score and immunohistochemical type II collagen staining). OA was surgically induced in the knee joint of rats by anterior cruciate/medial collateral ligament transection and medial meniscus resection (ACLT + MMx). Animals were divided into three groups: sham‐operated group (Sham), ACLT + MMx group without LIPUS (?LIPUS), and ACLT + MMx group with LIPUS (+LIPUS; 30 mW/cm², 20 min/day for 28 days). CTX‐II levels were elevated in both ?LIPUS and +LIPUS groups compared to that in the Sham group after the operation, but there was no significant difference between +LIPUS and ?LIPUS groups, suggesting that LIPUS does not affect the degradation of type II collagen in this model. In contrast, CPII was significantly increased in +LIPUS group compared to ?LIPUS and Sham. Moreover, histological damage on the cartilage (Mankin score) was ameliorated by LIPUS, and type II collagen was immunohistochemically increased by LIPUS in the cartilage of an OA model. Of interest, mRNA expression of type II collagen was enhanced by LIPUS in chondrocytes. Together these observations suggest that LIPUS is likely to increase the type II collagen synthesis in articular cartilage, possibly via the activation of chondrocytes and induction of type II collagen mRNA expression, thereby exhibiting chondroprotective action in a rat OA model. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:361–369, 2010     

Abnormal human chondrocyte morphology is related to increased levels of cell‐associated IL‐1β and disruption to pericellular collagen type VI

Early osteoarthritis (OA) is poorly understood, but abnormal chondrocyte morphology might be important. We studied IL‐1β and pericellular collagen type VI in morphologically normal and abnormal chondrocytes. In situ chondrocytes within explants from nondegenerate (grade 0/1) areas of human tibial plateaus (n = 21) were fluorescently labeled and visualized [2‐photon laser scanning microscopy (2PLSM)]. Normal chondrocytes exhibited a “smooth” membrane surface, whereas abnormal cells were defined as demonstrating ≥1 cytoplasmic process. Abnormal chondrocytes were further classified by number and average length of cytoplasmic processes/cell. IL‐1β or collagen type VI associated with single chondrocytes were visualized by fluorescence immuno‐histochemistry and confocal laser scanning microscopy (CLSM). Fluorescence was quantified as the number of positive voxels (i.e., 3D pixels with fluorescence above baseline)/cell. IL‐1β‐associated fluorescence increased between normal and all abnormal cells in the superficial (99.7 ± 29.8 [11 (72)] vs. 784 ± 382 [15 (132)]; p = 0.04, positive voxels/cell) and deep zones (66.5 ± 29.4 [9 (64)] vs. 795 ± 224 [9 (56)]; p = 0.006). There was a correlation (r² = 0.988) between the number of processes/cell (0–5) and IL‐1β, and an increase particularly with short processes (≤5 µm; p = 0.022). Collagen type VI coverage and thickness decreased (p < 0.001 and p = 0.005, respectively) with development of processes. Abnormal chondrocytes in macroscopically nondegenerate cartilage demonstrated a marked increase in IL‐1β and loss of pericellular type VI collagen, changes that could lead to cartilage degeneration. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1507–1514, 2010     

Spatial organization of types I and II collagen in the canine meniscus.

The meniscus of the knee joint is a fibrocartilage mainly composed of type I collagen and smaller amounts of type II collagen. The distribution of type II collagen in the canine meniscus and its spatial relationship to type I collagen was examined by immunohistochemistry and confocal microscopy. Dorsal and coronal slices of the mid-section of medial and lateral menisci from the knee joints of skeletally mature dogs were predigested with Streptomyces hyaluronate lyase and bacterial Protease enzyme XXIV. Monoclonal antibodies against type I collagen (CP17L) and type II collagen (II-II6B3) and an anti-type II collagen polyclonal antibody (AB759) were employed. The staining for type II collagen in the extracellular matrix of hyaline articular cartilage was diffuse without any identifiable spatial organization. In striking contrast, type II collagen in the fibrocartilage of the meniscus stained as an organized network. Type II collagen was distributed throughout the meniscus with the exception of the outer zone containing the blood vessels. Coronal and dorsal staining of the meniscus showed bundles of circumferential fibrils of type I that colocalized with type II collagen in specific sites. These bundles were enwrapped in a second organizational fibrillar system of types I and II collagen that also colocalized. Bundles of circumferential fibrils appeared in cross-section in coronal sections as dots within the interstitial spaces framed by the network of types I and II collagen of the second system. Confocal overlays showed that types I and II collagens were superimposed, suggesting a close spatial proximity between the two collagens. The cells were confined to the types I and II collagen fibrils that enwrapped the bundles. A striking feature of the radial tie fibers was patches of type II collagen without colocalized type I collagen. Our study reveals a unique network of type II collagen in fibrocartilage of the meniscus that serves as a morphological distinction between fibro- and hyaline cartilage.     

Articular cartilage repair with recombinant human type II collagen/polylactide scaffold in a preliminary porcine study

The purpose of this study was to investigate the potential of a novel recombinant human type II collagen/polylactide scaffold (rhCo‐PLA) in the repair of full‐thickness cartilage lesions with autologous chondrocyte implantation technique (ACI). The forming repair tissue was compared to spontaneous healing (spontaneous) and repair with a commercial porcine type I/III collagen membrane (pCo). Domestic pigs (4‐month‐old, n = 20) were randomized into three study groups and a circular full‐thickness chondral lesion with a diameter of 8 mm was created in the right medial femoral condyle. After 3 weeks, the chondral lesions were repaired with either rhCo‐PLA or pCo together with autologous chondrocytes, or the lesion was only debrided and left untreated for spontaneous repair. The repair tissue was evaluated 4 months after the second operation. Hyaline cartilage formed most frequently in the rhCo‐PLA treatment group. Biomechanically, there was a trend that both treatment groups resulted in better repair tissue than spontaneous healing. Adverse subchondral bone reactions developed less frequently in the spontaneous group (40%) and the rhCo‐PLA treated group (50%) than in the pCo control group (100%). However, no statistically significant differences were found between the groups. The novel rhCo‐PLA biomaterial showed promising results in this proof‐of‐concept study, but further studies will be needed in order to determine its effectiveness in articular cartilage repair. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:745–753, 2016.     

Angiotensin II further enhances type IV collagen production stimulated by platelet-derived growth factor and fibroblast growth factor-2 in cultured human mesangial cells

SUMMARY: Angiotensin II (Ang II) is considered to play a role in the development of glomerulosclerosis, which is characterized by excessive accumulation of mesangial matrix after mesangial cell proliferation. We have reported that platelet-derived growth factor (PDGF) and fibroblast growth factor-2 (FGF-2) stimulate type IV collagen production by cultured human mesangial cells (HMC). Although Ang II is well known to have a mitogenic effect in various kinds of cells, its role in the production of extracellular matrix is still undetermined. This study was designed to examine the effect of Ang II and its interaction with PDGF and FGF-2 in type IV collagen production by HMC. Cultured HMC were incubated with Ang II with or without PDGF or FGF-2 for 72 h and type IV collagen, fibronectin and laminin in the cell supernatants were measured. Ang II (10⁻⁶–10⁻⁸) itself did not change the production of type IV collagen, fibronectin, laminin and transforming growth factor-β. PDGF and FGF-2 enhanced type IV production, although they did not stimulate the production of fibronectin and laminin. Ang II further increased the stimulating effect of PDGF and FGF-2 in type IV collagen production in a dose-dependent manner. This effect of Ang II was completely blocked by Ang II type I receptor antagonist (Losartan). These data imply that Ang II is a potent stimulator of type IV collagen production by HMC in the presence of growth factors.     

E‐cadherin upregulates expression of matrix macromolecules aggrecan and collagen II in the intervertebral disc cells through activation of the intracellular BMP‐Smad1/5 pathway

E‐cadherin is a transmembrane protein that mediates cell–cell adhesion and cell–matrix interaction. Although the E‐cadherin has been shown to mediate a broad‐ranging cellular signals and functions, its effects on matrix metabolism of intervertebral discs (IVDs) are unknown. In this study, we investigated the effects of E‐cadherin on IVD matrix synthesis using pharmacological and molecular biology methods. We showed that high levels of the E‐cadherin are expressed in rabbits IVD cells. Our study indicates that the ectopic expression of E‐cadherin can stimulate matrix anabolism of the IVD cells, which was evidenced by increased expression of the matrix macromolecules aggrecan and collagen II. We found that E‐cadherin induces the expression of BMP‐4 and BMP‐7 genes and enhances Smad1/5 phosphorylation. Blocking BMP activity uses noggin suppressed E‐cadherin‐mediated upregulation of aggrecan and collagen II. Moreover, inhibition of Smad1/5 phosphorylation by dorsomorphin significantly repressed the E‐cadherin induced expression of aggrecan and collagen II at the both mRNA and protein levels. Together this study demonstrates that the E‐cadherin stimulates the synthesis of IVD matrix macromolecules aggrecan and collagen II through the induction of BMP genes and enhancement of the Smad1/5 phosphorylation. Thus E‐cadherin may have value in the treatment of degenerated discs. © 2012 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 30:1746–1752, 2012     

Similar rate of progression in the predialysis phase in type I and type II diabetes mellitus

Progression of diabetic nephropathy from the stage of macroproteinuriawith near-normal renal function until start of dialysis wascompared in 16 patients with type I and 16 patients with typeII diabetes mellitus. The mean creatinine clearance at the beginningof the study was 89±13 ml/min/l.73 m² in patients withtype I and 81±6 ml/min/1.73 m in those with type II diabetes.Dialysis was started after a mean interval of 77(44–133)months, when creatinine clearance had decreased to 8 ±2 ml/min/1.73 m² in type I diabetic patients. The respectivefigures for type II diabetic patients were 81(40–124)months and 7±2 ml/min/1.73 m² The mean rate of decreasein creatinine clearance was 1.05 ± 0.45 ml/min/monthin type I and 0.91 ± 0.41 ml/min/month in type II diabetes.The mean rate of decrease was 1.46±0.30 ml/min/monthin type I diabetic patients with a systolic BP> 160 mmHgversus 0.80±0.42 ml/mm/month with <160 minHg (P>0.01).In the type II diabetics the respective figures were 1.38±0.40m1/min/monthversus 0.78 ± 0.15 ml/min/month (P>0.01). During theobservation period the prevalence of coronary heart diseaseincreased from 6 to 50% in type I and from 31 to 87% in typeII diabetes. In conclusion, the rate of progression of diabeticnephro pathy during the predialytic phase is similar in typeI and type II diabetes; BP adversely affects the rate of progressionto the same extent in both groups.     

Biomarkers,type II collagen,glucosamine and chondroitin sulfate in osteoarthritis follow-up: the “Magenta osteoarthritis study”

Background  The purpose of the present study was to determine relationship between disease activity, systemic markers of cartilage degradation, urinary C-terminal cross-linking telopeptides of type II collagen (uCTX-II), and bone degradation, urinary C-terminal cross-linking telopeptides of type I collagen (uCTX-I), structural progression of osteoarthritis (OA) and potential therapeutic efficacy of type II collagen (COLLII) in combination with glucosamine and chondroitin sulfate (GC). Materials and methods  An observational retrospective study, 1-year follow-up, on 104 patients with OA (nodular osteoarthritis of the hand, erosive osteoarthritis of the hand, EOA, osteoarthritis of the knee or hip) who were treated with GC or glucosamine, chondroitin sulfate and collagen type II (GCC). The following information was collected at entry: demographics, BMI, characteristics of OA, patient global assessment (VAS), C-terminal cross-linking telopeptides of collagen types I (uCTX-I) and II (uCTX-II) and radiographs. After 6 months: VAS, uCTX-I and uCTX-II. After 1 year: VAS, uCTX-I, uCTX-II and radiographs. Results  After 6 months and 1 year of treatment VAS, uCTX-I and uCTX-II mean values were significantly lower than the baseline. 57 were treated with GCC and 47 with GC. The group that received GCC showed a similar VAS mean value after 6 months and 1 year when compared with the group treated with GC. uCTX-I and uCTX-II mean level was lower in the group treated with GCC (P < 0.05). Radiological score (Kellgren and Lawrence summarized score for hands) after 1 year showed a reduced progression compared to the baseline in the hand osteoarthritis group, especially after GCC treatment (P < 0.05). Finally, uCTX-I has better correlation with radiological score and with GC in the EOA subgroup (Pearson index: R = 0.44). Conclusions  (a) uCTX-I and uCTX-II proved to be useful biomarkers in OA monitoring; (b) uCTX-I is better correlated with hand EOA and could represent a potential further marker to assess the evolution of EOA bone damage; (c) GC slow down OA progression; (d) finally COLLII could represent a further protective factor in OA cartilage.     

Effects of the angiotensin II type 1 receptor antagonist candesartan,compared with angiotensin-converting enzyme inhibitors,on the urinary excretion of albumin and type IV collagen in patients with diabetic nephropathy

Background. We previously reported that the angiotensin II type 1 receptor antagonist candesartan was effective in reducing blood pressure and microalbuminuria in hypertensive patients with diabetic nephropathy after angiotensin-converting enzyme (ACE) inhibitors were replaced due to side effects. In the present study, the clinical effects of candesartan were investigated and compared with ACE inhibitors in patients with stage 2 or 3A diabetic nephropathy, mainly with respect to the effects on the urinary excretion of albumin and type IV collagen. Methods. Forty-nine patients (26 males/23 females) with diabetic nephropathy (stage 2 or 3A), including normotensive patients, were the study subjects. The patients were treated with either an ACE inhibitor (23 patients) or candesartan (26 patients) for 11 ± 3 months. The urinary excretion of albumin and urinary type IV collagen was measured. Results. Posttreatment blood pressure tended to decrease, but such a decrease did not reach a statistically significant level, nor did it show any intergroup difference. The urinary albumin excretion was positively correlated with pretreatment mean blood pressure and left ventricular mass index, but the urinary type IV collagen excretion did not show such correlations. The urinary albumin excretion decreased significantly after treatment to a similar extent in both groups, whereas the urinary type IV collagen excretion decreased significantly only in the candesartan group. Conclusion. It was revealed that ACE inhibitors and candesartan reduced urinary albumin excretion to a similar extent in patients with diabetic nephropathy. From the results of the present study, it is inferred that the renoprotective effect of candesartan in diabetic nephropathy may partially differ from that of ACE inhibitors.     

Changes in serum collagen markers,IGF‐I,and Knee joint laxity across the menstrual cycle

Variations in serum markers of collagen production (CICP) and degradation (ICTP), insulin‐like growth factor I (IGF‐I) and anterior knee laxity (AKL) were measured in 20 women [10 with spontaneous cycles (eumenorrheic), 10 using oral contraceptives] over 5 consecutive days at menses (M1–M5, 1st pill week), the initial estrogen rise near ovulation (O1–O5, 2nd pill week), the initial progesterone rise of the early luteal phase (EL1–EL5, 3rd pill week) and post‐progesterone peak of the late luteal phase (LL1–LL5, 4th pill week). ICTP was higher in oral contraceptive women (5.3 ± 1.7 vs. 3.7 ± 1.3 µg/L; p = 0.030), primarily during days near ovulation and the early luteal phase when concentrations decreased in eumenorrheic women (p = 0.04). IGF‐I concentrations increased during menses then decreased and remained lower during the early and late luteal phase in oral contraceptive women, resulting in lower concentrations compared to eumenorrheic women at EL2 and LL1 (p = 0.03). CICP decreased in early and late luteal days (p <0.01), and there was a trend toward lower concentrations in eumenorrheic versus oral contraceptive women (85.7 ± 35.7 ng/ml vs. 123.2 ± 49.8 ng/ml; p = 0.07). Lower CICP and greater IGF‐I concentrations predicted greater AKL across the 20 cycle days in both groups (R² = 0.310 and 0.400). Sex hormone concentration changes across the menstrual cycle are of sufficient magnitude to influence collagen metabolism, and may indirectly influence knee structure and function. © 2012 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 30:1405–1412, 2012     

Expression patterns of collagen types I and III in the capsule of a rat knee contracture model

Our objective was to determine the changes in expression of collagen types I and III in the capsule of a rat knee contracture model. The unilateral knee joints of adult male rats were rigidly immobilized at 150° of flexion using a rigid plastic plate and screws for 3 days, 1, 2, 4, 8, and 16 weeks (immobilized group). Sham‐operated animals had holes drilled in the femur and tibia with screws inserted without a plate (control group). The expression patterns of collagen types I and III in the anterior and posterior capsule were evaluated by in situ hybridization (ISH), quantitative real‐time polymerase chain reaction (qPCR), immunohistochemistry (IHC), and Western blotting (WB). Expressions of collagen types I and III were decreased after immobilization compared to the control group by ISH and qPCR. The expression was not changed after immobilization compared to the control group by IHC and WB. The expression of mRNA and protein levels of collagen types I and III were not increased after immobilization, which indicated that accumulation of the two types of collagen was not the etiology of joint contracture. Another process, such as capsule and synovial adhesions, may be one possible cause of joint contracture. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:315–321, 2010