The nutrition of the human meniscus: A computational analysis investigating the effect of vascular recession on tissue homeostasis

The meniscus is essential to the functioning of the knee, offering load support, congruency, lubrication, and protection to the underlying cartilage. Meniscus degeneration affects ∼35% of the population, and potentially leads to knee osteoarthritis. The etiology of meniscal degeneration remains to be elucidated, although many factors have been considered. However, the role of nutritional supply to meniscus cells in the pathogenesis of meniscus degeneration has been so far overlooked. Nutrients are delivered to meniscal cells through the surrounding synovial fluid and the blood vessels present in the outer region of the meniscus. During maturation, vascularization progressively recedes up to the outer 10% of the tissue, leaving the majority avascular. It has been hypothesized that vascular recession might significantly reduce the nutrient supply to cells, thus contributing to meniscus degeneration. The objective of this study was to evaluate the effect of vascular recession on nutrient levels available to meniscus cells. This was done by developing a novel computational model for meniscus homeostasis based on mixture theory. It was found that transvascular transport of nutrients in the vascularized region of the meniscus contributes to more than 40% of the glucose content in the core of the tissue. However, vascular recession does not significantly alter nutrient levels in the meniscus, reducing at most 5% of the nutrient content in the central portion of the tissue. Therefore, our analysis suggests that reduced vascularity is not likely a primary initiating source in tissue degeneration. However, it does feasibly play a key role in inability for self-repair, as seen clinically.     

Mesenchymal stem cells for enhancing biological healing after meniscal injuries

The meniscus is a semilunar fibrocartilage structure that plays important roles in maintaining normal knee biomechanics and function. The roles of the meniscus, including load distribution, force transmission, shock absorption, joint stability, lubrication, and proprioception, have been well established. Injury to the meniscus can disrupt overall joint stability and cause various symptoms including pain, swelling, giving-way, and locking. Unless treated properly, it can lead to early degeneration of the knee joint. Because meniscal injuries remain a significant challenge due to its low intrinsic healing potential, most notably in avascular and aneural inner two-thirds of the area, more efficient repair methods are needed. Mesenchymal stem cells (MSCs) have been investigated for their therapeutic potential in vitro and in vivo. Thus far, the application of MSCs, including bone marrow-derived, synovium-derived, and adipose-derived MSCs, has shown promising results in preclinical studies in different animal models. These preclinical studies could be categorized into intra-articular injection and tissue-engineered construct application according to delivery method. Despite promising results in preclinical studies, there is still a lack of clinical evidence. This review describes the basic knowledge, current treatment, and recent studies regarding the application of MSCs in treating meniscal injuries. Future directions for MSC-based approaches to enhance meniscal healing are suggested.     

Structured three-dimensional co-culture of mesenchymal stem cells with meniscus cells promotes meniscal phenotype without hypertrophy

Menisci play a crucial role in weight distribution, load bearing, shock absorption, lubrication, and nutrition of articular cartilage within the knee joint. Damage to the meniscus typically does not heal spontaneously due to its partial avascular nature. Partial or complete meniscectomy is a common clinical treatment of the defective meniscus. However, this procedure ultimately leads to osteoarthritis due to increased mechanical stress to the articular cartilage. Meniscus tissue engineering offers a promising solution for partial or complete meniscus deficiency. Mesenchymal stem cells (MSC) have the potential to differentiate into meniscal fibrochondrocyte as well as deliver trophic effects to the differentiated cells. This study tested the feasibility of using MSC co-cultured with mature meniscal cells (MC) for meniscus tissue engineering. Structured cell pellets were created using MC and MSC at varying ratios (100:0, 75:25, 50:50, 25:75, and 0:100) and cultured with or without transforming growth factor-beta 3 supplemented chondrogenic media for 21 days. The meniscal and hypertrophic gene expression, gross appearance and structure of the pellets, meniscus extracellular matrix (ECM), histology and immunohistochemistry of proteoglycan and collagen were evaluated. Co-culture of MC with MSC at 75:25 demonstrated highest levels of collagen type I and glycosaminoglycans (GAG) production, as well as the lowest levels of hypertrophic genes, such as COL10A1 and MMP13. All co-culture conditions showed better meniscus ECM production and hypertrophic inhibition as compared to MSC culture alone. The collagen fiber bundles observed in the co-cultures are important to produce heterogenic ECM structure of meniscus. In conclusion, co-culturing MC and MSC is a feasible and efficient approach to engineer meniscus tissue with enhanced ECM production without hypertrophy.     

Meniscus maturation in the swine model: changes occurring along with anterior to posterior and medial to lateral aspect during growth

The meniscus plays important roles in knee function and mechanics and is characterized by a heterogeneous matrix composition. The changes in meniscus vascularization observed during growth suggest that the tissue‐specific composition may be the result of a maturation process. This study has the aim to characterize the structural and biochemical variations that occur in the swine meniscus with age. To this purpose, menisci were collected from young and adult pigs and divided into different zones. In study 1, both lateral and medial menisci were divided into the anterior horn, the body and the posterior horn for the evaluation of glycosaminoglycans (GAGs), collagen 1 and 2 content. In study 2, the menisci were sectioned into the inner, the intermediate and the outer zones to determine the variations in the cell phenotype along with the inner–outer direction, through gene expression analysis. According to the results, the swine meniscus is characterized by an increasing enrichment in the cartilaginous component with age, with an increasing deposition in the anterior horn (GAGs and collagen 2; P < 0.01 both); moreover, this cartilaginous matrix strongly increases in the inner avascular and intermediate zone, as a consequence of a specific differentiation of meniscal cells towards a cartilaginous phenotype (collagen 2, P < 0.01). The obtained data add new information on the changes that accompany meniscus maturation, suggesting a specific response of meniscal cells to the regional mechanical stimuli in the knee joint.     

Regional variation in the mechanical role of knee meniscus glycosaminoglycans

High compressive properties of cartilaginous tissues are commonly attributed to the sulfated glycosaminoglycan (GAG) fraction of the extracellular matrix (ECM), but this relationship has not been directly measured in the knee meniscus, which shows regional variation in GAG content. In this study, biopsies from each meniscus region (outer, middle, and inner) were either subjected to chondroitinase ABC (CABC) to remove all sulfated GAGs or not. Compressive testing revealed that GAG depletion in the inner and middle meniscus regions caused a significant decrease in modulus of relaxation (58% and 41% decreases, respectively, at 20% strain), and all regions exhibited a significant decrease in viscosity (outer: 29%; middle: 58%; inner: 62% decrease). Tensile properties following CABC treatment were unaffected for outer and middle meniscus specimens, but the inner meniscus displayed significant increases in Young's modulus (41% increase) and ultimate tensile stress (40% increase) following GAG depletion. These findings suggest that, in the outer meniscus, GAGs contribute to increasing tissue viscosity, whereas in the middle and inner meniscus, where GAGs are most abundant, these molecules also enhance the tissue's ability to withstand compressive loads. GAGs in the inner meniscus also contribute to reducing the circumferential tensile properties of the tissue, perhaps due to the pre-stress on the collagen network from increased hydration of the ECM. Understanding the mechanical role of GAGs in each region of the knee meniscus is important for understanding meniscus structure-function relationships and creating design criteria for functional meniscus tissue engineering efforts.     

Effects of hepatocyte growth factor and platelet-derived growth factor on the repair of meniscal defects in vitro

Summary Injuries to the avascular region of the meniscus occur frequently and may be difficult to repair. This study was designed to determine whether growth factors could diffuse from a collagen sponge or a collagen gel into meniscal tissue and stimulate healing of defects using an in vitro model. The diffusion of platelet-derived growth factor (PDGF) from the collagen carriers into the medium was rapid with approximately 50% being released from the collagen sponge within the first hour. After 5 d of incubation, 8% of the PDGF was present in the meniscus, 11% in the collagen sponge, and 62% had been released into the medium. Similar results were obtained when a collagen gel was used as a carrier. Histological evaluation of the meniscal explants after 2 wk in culture revealed extensive proteoglycan staining in the areas surrounding defects treated with either hepatocyte growth factor (HGF) or PDGF compared with controls without growth factor. The HGF-PDGF treatment resulted in alignment and migration of meniscal cells toward the defect, which was not observed in untreated controls. At 3–7 d, increased number of cells were observed in defects treated with collagen gels (but not the sponge) with PDGF-HGF. At 4 wk, combined HGF-PDGF treatment resulted in the formation of tissue with birefringence by polarized microscopy, suggestive of organized collagen. The data suggest that use of specific PDGF-HGF may enhance the repair of meniscal injuries.     

An in vitro model to assess mechanisms and efficacy of a cellular conduit for treatment of avascular meniscal injuries

Tears in the avascular portion of the knee meniscus are commonplace and are frequently incapable of healing spontaneously. Delivery of synovial cells from the meniscal periphery to avascular injuries can result in an effective healing response but is difficult to accomplish surgically. This report describes the development of a novel in vitro model comprised of three-dimensionally cultured cells in agarose used to assess the proof of concept that a cellular conduit device could be used to facilitate the delivery of synovial fibroblasts from a cell source to a remote acellular recipient site. The results indicate that synovial fibroblasts are capable of migrating through a cellular conduit more optimally than a created trephined channel over a clinically relevant distance in response to a chemotactic gradient. This model proved to be a reliable way to assess fibroblast-like synoviocyte migration in a clinically relevant fashion for application to avascular meniscal tear healing methodologies, and provided mechanistic information regarding the successful in vivo testing of this specific biomedical device.     

Integrative repair of the meniscus: lessons from in vitro studies

Current therapies for meniscal injury seek to preserve and repair damaged tissue since loss of meniscal tissue is associated with degenerative changes in the joint, ultimately leading to osteoarthritis (OA). After a meniscal tear, the difficulty of integrating juxtaposed meniscal surfaces continues to be an obstacle. In order to determine the local factors that are necessary for successful tissue repair, previous studies have developed in vitro model systems that allow both biological and quantitative biomechanical measures of meniscus repair. Many studies have shown the importance of individual factors in meniscus metabolism, but there is a complex interplay among a variety of factors that influence meniscal healing, including inflammatory cytokines, growth factors, mechanical loading, and zonal differences in cell and tissue properties. In particular, the upregulation of inflammatory cytokines following joint injury appears to have significant catabolic influences on meniscal cell metabolic activity that must be overcome in order to promote repair. In the presence of inflammatory cytokines, such as interleukin-1 (IL-1) or tumor necrosis factor alpha (TNF-alpha), intrinsic meniscal repair in vitro is significantly inhibited. While anabolic growth factors, such as transforming growth factor-beta1 (TGF-beta1), enhance meniscal repair, they cannot completely overcome the IL-1-mediated inhibition of repair. The mechanisms by which these mediators influence meniscal repair, and their interactions with other factors in the microenvironment, such as mechanical loading, remain to be determined. Future studies must address these complex interactions during meniscal healing to ultimately enhance meniscal repair.     

Interleukin-1, tumor necrosis factor-alpha,and transforming growth factor-beta 1 and integrative meniscal repair: influences on meniscal cell proliferation and migration

Introduction  

Interleukin-1 (IL-1) and tumor necrosis factor-α (TNF-α) are up-regulated in injured and osteoarthritic knee joints. IL-1 and TNF-α inhibit integrative meniscal repair; however, the mechanisms by which this inhibition occurs are not fully understood. Transforming growth factor-β1 (TGF-β1) increases meniscal cell proliferation and accumulation, and enhances integrative meniscal repair. An improved understanding of the mechanisms modulating meniscal cell proliferation and migration will help to improve approaches for enhancing intrinsic or tissue-engineered repair of the meniscus. The goal of this study was to examine the hypothesis that IL-1 and TNF-α suppress, while TGF-β1 enhances, cellular proliferation and migration in cell and tissue models of meniscal repair.     

Meniscal pathology - the evidence for treatment

Whilst arthroscopic surgery for the treatment of meniscal tears is the most commonly performed orthopaedic surgery, meniscal tears at the knee are frequently identified on magnetic resonance imaging in adults with and without knee pain. The evidence for arthroscopic treatment of meniscal tears is controversial and lacks a supporting evidence base; it may be no more efficacious than conservative therapies. Surgical approaches to the treatment of meniscal pathology can be broadly categorised into those in which partial menisectomy or repair are performed. This review highlights that the major factor determining the choice of operative approach is age: meniscal repair is performed exclusively on younger populations, while older populations are subject to partial menisectomy procedures. This is probably because the meniscus is less amenable to repair in the older population where other degenerative changes co-exist. In middle-aged to older adults, arthroscopic partial menisectomy (APM) may treat the meniscus tear, but does not address the degenerative whole organ disease of knee osteoarthritis. Thus far, there is no convincing evidence that operative approaches are superior to conservative measures as the first-line treatment of older people with knee pain and meniscal tears. However, in two randomised controlled trials (RCTs) approximately one-third of subjects in the exercise groups had persisting knee pain with some evidence of improvement following APM, although the characteristics of this subgroup are unclear. From the available data, a first-line trial of conservative therapy, which includes weight loss, is recommended for the treatment of degenerative meniscal tears in older adults. The exception to this may be when mechanical symptoms, such as knee locking, predominate. Although requiring corroboration by RCTs, there is accumulating evidence from cohort studies and case series that meniscal repair rather than APM may improve function and reduce the long-term risk of knee osteoarthritis in young adults. There is no clear evidence from RCTs that one surgical method of meniscal repair is superior to another.     

Age‐related modulation of angiogenesis‐regulating factors in the swine meniscus

An in‐depth knowledge of the native meniscus morphology and biomechanics in its different areas is essential to develop an engineered tissue. Meniscus is characterized by a great regional variation in extracellular matrix components and in vascularization. Then, the aim of this work was to characterize the expression of factors involved in angiogenesis in different areas during meniscus maturation in pigs. The menisci were removed from the knee joints of neonatal, young and adult pigs, and they were divided into the inner, intermediate and outer areas. Vascular characterization and meniscal maturation were evaluated by immunohistochemistry and Western blot analysis. In particular, expression of the angiogenic factor Vascular Endothelial Growth Factor (VEGF) and the anti‐angiogenic marker Endostatin (ENDO) was analysed, as well as the vascular endothelial cadherin (Ve‐CAD). In addition, expression of Collagen II (COLL II) and SOX9 was examined, as markers of the fibro‐cartilaginous differentiation. Expression of VEGF and Ve‐CAD had a similar pattern in all animals, with a significant increase from the inner to the outer part of the meniscus. Pooling the zones, expression of both proteins was significantly higher in the neonatal meniscus than in young and adult menisci. Conversely, the young meniscus revealed a significantly higher expression of ENDO compared to the neonatal and adult ones. Analysis of tissue maturation markers showed an increase in COLL II and a decrease in SOX9 expression with age. These preliminary data highlight some of the changes that occur in the swine meniscus during growth, in particular the ensemble of regulatory factors involved in angiogenesis.     

Finite Element Modeling Predictions of Region-specific Cell-matrix Mechanics in the Meniscus

The knee meniscus exhibits significant spatial variations in biochemical composition and cell morphology that reflect distinct phenotypes of cells located in the radial inner and outer regions. Associated with these cell phenotypes is a spatially heterogeneous microstructure and mechanical environment with the innermost regions experiencing higher fluid pressures and lower tensile strains than the outer regions. It is presently unknown, however, how meniscus tissue mechanics correlate with the local micromechanical environment of cells. In this study, theoretical models were developed to study mechanics of inner and outer meniscus cells with varying geometries. The results for an applied biaxial strain predict significant regional differences in the cellular mechanical environment with evidence of tensile strains along the collagen fiber direction of ~0.07 for the rounded inner cells, as compared to levels of 0.02–0.04 for the elongated outer meniscus cells. The results demonstrate an important mechanical role of extracellular matrix anisotropy and cell morphology in regulating the region-specific micromechanics of meniscus cells, that may further play a role in modulating cellular responses to mechanical stimuli.     

Mechanical injury of explants from the articulating surface of the inner meniscus

Knee osteoarthritis is accelerated by damage to the meniscus, a fibrocartilage tissue that assists in load transmission. However, little is known about the mechanical or cellular response of the meniscus to injurious overloading. Here, in vitro studies explored injury to meniscal explants using a compressive overloading protocol that has been well characterized for articular cartilage. Cartilage samples were processed in parallel as a reference to the extensive literature on cartilage injury. Injured meniscal explants showed extensive cell death at the articulating surface but no gross tissue damage, while similar conditions of peak stress and strain resulted in cartilage surface fissures and cell death consistent with moderate overloading. Post-injury gene expression in meniscal explants indicated a decrease in seven of the nine catabolic and pro-inflammatory molecules surveyed, while cartilage experienced a downregulation in ADAMTS-5 and TNF-α only. These data demonstrated a resiliency of the meniscus to injury, and that an acute increase in catabolic activities is not necessarily a consequence of mechanical overloading.     

Stresses and strains in the medial meniscus of an ACL deficient knee under anterior loading: a finite element analysis with image-based experimental validation

The menisci are believed to play a stabilizing role in the ACL-deficient knee, and are known to be at risk for degradation in the chronically unstable knee. Much of our understanding of this behavior is based on ex vivo experiments or clinical studies in which we must infer the function of the menisci from external measures of knee motion. More recently, studies using magnetic resonance (MR) imaging have provided more clear visualization of the motion and deformation of the menisci within the tibio-femoral articulation. In this study, we used such images to generate a finite element model of the medial compartment of an ACL-deficient knee to reproduce the meniscal position under anterior loads of 45, 76, and 107 N. Comparisons of the model predictions to boundaries digitized from images acquired in the loaded states demonstrated general agreement, with errors localized to the anterior and posterior regions of the meniscus, areas in which large shear stresses were present. Our model results suggest that further attention is needed to characterize material properties of the peripheral and horn attachments. Although overall translation of the meniscus was predicted well, the changes in curvature and distortion of the meniscus in the posterior region were not captured by the model, suggesting the need for refinement of meniscal tissue properties.     

Is the meniscus of the knee joint a fibrocartilage?

A histological analysis of the structure of intact knee joint menisci was carried out in adult dogs. By means of specific histochemical methods for the connective tissue and cartilage, it was found that the meniscus as a whole does not have a unique structure. The anterior and posterior horns are populated by round chondroid cells encircled by abundant interstitial substance and branched wavy connective fibers; blood vessels are present. The outer third of the meniscus is constituted of cross bundles of connective fibers, fibrocytes and spindle-like areas of loose connective tissue with blood vessels. The inner avascular two thirds of the meniscus are filled with parallel circumferentially oriented fascicles of connective fibers, ovally elongated chondroid cells, and a small quantity of chondroid interstitial substance. In some menisci, in the inner two thirds of the body, there are isles of typical cartilage, which show metachromasia of the beta type and rarely of the gamma type. The occurrence and way of the manifestation of cartilage are of an individual character. The structural duality of the knee meniscus is accounted for by its functional duality manifested in offering resistance to the forces of traction and pressure, the latter ones favoring the process of evolution of tissue from connective, through chondroid, to cartilaginous.     

Regional differences in prostaglandin E2 and nitric oxide production in the knee meniscus in response to dynamic compression

Injury or loss of the knee meniscus is associated with altered joint stresses that lead to progressive joint degeneration. The goal of this study was to determine if dynamic mechanical compression influences the production of inflammatory mediators by meniscal cells. Dynamic compression increased prostaglandin E2 (PGE(2)) and nitric oxide (NO) production over a range of stress magnitudes (0.0125-0.5 MPa) in a manner that depended on stress magnitude and zone of tissue origin. Inner zone explants showed greater increases in PGE(2) and NO production as compared to outer zone explants. Meniscal tissue expressed NOS2 and NOS3 protein, but not NOS1. Mechanically induced NO production was blocked by NOS inhibitors, and the non-selective NOS inhibitor L-NMMA augmented PGE(2) production in the outer zone only. These findings suggest that the meniscus may serve as an intra-articular source of pro-inflammatory mediators, and that alterations in the magnitude or distribution of joint loading could significantly influence the production of these mediators in vivo.     

Gene therapy and tissue engineering for sports medicine

Sports injuries usually involve tissues that display a limited capacity for healing. The treatment of sports injuries has improved over the past 10 to 20 years through sophisticated rehabilitation programs, novel operative techniques, and advances in the field of biomechanical research. Despite this considerable progress, no optimal solution has been found for treatment of various sports-related injuries, including muscle injuries, ligament and tendon ruptures, central meniscal tears, cartilage lesions, and delayed bone fracture healing. New biological approaches focus on the treatment of these injuries with growth factors to stimulate and hasten the healing process. Gene therapy using the transfer of defined genes encoding therapeutic proteins represents a promising way to efficiently deliver suitable growth factors into the injured tissue. Tissue engineering, which may eventually be combined with gene therapy, may potentially result in the creation of tissues or scaffolds for regeneration of tissue defects following trauma. In this article we will discuss why gene therapy and tissue engineering are becoming increasingly important in modern orthopaedic sports medicine practice. We then will review recent research achievements in the area of gene therapy and tissue engineering for sports-related injuries, and highlight the potential clinical applications of this technology in the treatment of patients with musculoskeletal problems following sports-related injuries.     

Proteoglycans contribute locally to swelling,but globally to compressive mechanics,in intact cervine medial meniscus

Loss of charged proteoglycans in the knee meniscus, which aid in the support of compressive loads by entraining water, is an effect of degeneration and is often associated with osteoarthritis. In healthy menisci, proteoglycan content is highest in the inner white zone and decreases towards the peripheral red zone. We hypothesized that loss of proteoglycans would reduce both osmotic swelling and compressive stiffness, spatially localized to the avascular white zone of the meniscus. This hypothesis was tested by targeted enzymatic digestion of proteoglycans using hyaluronidase in intact cervine medial menisci. Mechanics were quantified by creep indentation on the femoral surface. Osmotic swelling changes were assessed by measuring collagen fiber crimp period in the radial-axial plane in the lamellar layer along both the tibial and femoral contacting surfaces. All measurements were made in the inner, middle, and outer zones of the anterior, central, and posterior regions. Mechanical measurements showed variation in creep behavior with anatomical location, along with spatially uniform decreases in viscosity (average of 21%) and creep stiffness (average of 15%) with hyaluronidase treatment. Lamellar collagen crimp period was significantly decreased (average of 27%) by hyaluronidase, indicating a decrease in osmotic swelling, with the largest decreases seen in locations with the highest proteoglycan content. Taken together, these results suggest that while proteoglycans have localized effects on meniscus swelling, the resulting effect on compressive properties is distributed throughout the tissue.     

Comparative spatial and temporal localisation of perlecan, aggrecan and type I, II and IV collagen in the ovine meniscus: an ageing study

This is the first study to immunolocalise perlecan in meniscal tissues and to demonstrate how its localisation varied with ageing relative to aggrecan and type I, II and IV collagen. Perlecan was present in the middle and inner meniscal zones where it was expressed by cells of an oval or rounded morphology. Unlike the other components visualised in this study, perlecan was strongly cell associated and its levels fell significantly with age onset and cell number decline. The peripheral outer meniscal zones displayed very little perlecan staining other than in small blood vessels. Picrosirius red staining viewed under polarised light strongly delineated complex arrangements of slender discrete randomly oriented collagen fibre bundles as well as transverse, thick, strongly oriented, collagen tie bundles in the middle and outer meniscal zones. The collagen fibres demarcated areas of the meniscus which were rich in anionic toluidine blue positive proteoglycans; immunolocalisations confirmed the presence of aggrecan and perlecan. When meniscal sections were examined macroscopically, type II collagen localisation in the inner meniscal zone was readily evident in the 2- to 7-day-old specimens; this became more disperse in the older meniscal specimens. Type I collagen had a widespread distribution in all meniscal zones at all time points. Type IV collagen was strongly associated with blood vessels in the 2- to 7-day-old meniscal specimens but was virtually undetectable at the later time points (>7 month).     

Human meniscus cells express hypoxia inducible factor-1alpha and increased SOX9 in response to low oxygen tension in cell aggregate culture

In previous work we demonstrated that the matrix-forming phenotype of cultured human cells from whole meniscus was enhanced by hypoxia (5% oxygen). Because the meniscus contains an inner region that is devoid of vasculature and an outer vascular region, here we investigate, by gene expression analysis, the separate responses of cells isolated from the inner and outer meniscus to lowered oxygen, and compared it with the response of articular chondrocytes. In aggregate culture of outer meniscus cells, hypoxia (5% oxygen) increased the expression of type II collagen and SOX9 (Sry-related HMG box-9), and decreased the expression of type I collagen. In contrast, with inner meniscus cells, there was no increase in SOX9, but type II collagen and type I collagen increased. The articular chondrocytes exhibited little response to 5% oxygen in aggregate culture, with no significant differences in the expression of these matrix genes and SOX9. In both aggregate cultures of outer and inner meniscus cells, but not in chondrocytes, there was increased expression of collagen prolyl 4-hydroxylase (P4H)alpha(I) in response to 5% oxygen, and this hypoxia-induced expression of P4H alpha(I) was blocked in monolayer cultures of meniscus cells by the hypoxia-inducible factor (HIF)-1alpha inhibitor (YC-1). In fresh tissue from the outer and inner meniscus, the levels of expression of the HIF-1alpha gene and downstream target genes (namely, those encoding P4H alpha(I) and HIF prolyl 4-hydroxylase) were significantly higher in the inner meniscus than in the outer meniscus. Thus, this study revealed that inner meniscus cells were less responsive to 5% oxygen tension than were outer meniscus cells, and they were both more sensitive than articular chondrocytes from a similar joint. These results suggest that the vasculature and greater oxygen tension in the outer meniscus may help to suppress cartilage-like matrix formation.