Non-destructive electromechanical assessment (Arthro-BST) of human articular cartilage correlates with histological scores and biomechanical properties.

OBJECTIVE: The hand-held Arthro-BST™ device is used to map electromechanical properties of articular cartilage. The purpose of the study was to evaluate correlation of electromechanical properties with histological, biochemical and biomechanical properties of cartilage. METHOD: Electromechanical properties (quantitative parameter (QP)) of eight human distal femurs were mapped manually ex vivo using the Arthro-BST (1 measure/site, 5 s/measure, 3209 sites). Osteochondral cores were then harvested from different areas on the femurs and assessed with the Mankin histological score. Prior to histoprocessing, cores were tested in unconfined compression. A subset of the cores was analyzed with polarized light microscopy (PLM) to assess collagen structure. Biochemical assays were done on additional cores to obtain water content and glycosaminoglycan (GAG) content. The QP corresponding to each core was calculated by averaging all QPs collected within 6 mm of the core center. RESULTS: The electromechanical QP correlated strongly with both the Mankin score and the PLM score (r = 0.73, P < 0.0001 and r = -0.70, P < 0.0001 respectively) thus accurately reflecting tissue quality and collagen architecture. Electromechanical QP also correlated strongly with biomechanical properties including fibril modulus (r = -0.76, P < 0.0001), matrix modulus (r = -0.69, P < 0.0001), and log of permeability (r = 0.72, P < 0.0001). The QP correlated weakly with GAG per wet weight and with water content (r = -0.50, P < 0.0003 and r = 0.39, P < 0.006 respectively). CONCLUSION: Non-destructive electromechanical QP measurements correlate strongly with histological scores and biomechanical parameters providing a rapid and reliable assessment of articular cartilage quality.

Stereological analysis of subchondral angiogenesis induced by chitosan and coagulation factors in microdrilled articular cartilage defects.

OBJECTIVE: Cartilage repair elicited by bone marrow stimulation can be enhanced by a chitosan-glycerol phosphate (GP)/blood implant, through mechanisms involving therapeutic inflammatory angiogenesis. The implant is formed by in situ coagulation, which can be accelerated by adding coagulation factors. We hypothesized that coagulation factors enhance acute subchondral angiogenesis in repairing drilled defects. DESIGN: Full-thickness cartilage defects were created bilaterally in 12 skeletally mature rabbit knee trochlea, microdrilled, then allowed to bleed as a control (N = 6) or treated with chitosan-GP/blood implant (N = 6), or implant solidified with thrombin (IIa), tissue factor (TF) with recombinant human factor VIIa (rhFVIIa), or rhFVIIa alone (N = 4 each condition). At 3 weeks post-operative, quantitative stereology was used to obtain blood vessel length (L(V)), surface (S(V)), and volume (V(V)) density at systematic depths in two microdrill holes per defect. Collagen type I, type II and glycosaminoglycan (GAG) percent stain in non-mineralized repair tissue were analysed by histomorphometry. RESULTS: All drill holes were healing, and showed a depth-dependent increase in granulation tissue blood vessel density (Lv, Sv, and Vv, P < 0.005). Residual chitosan implant locally suppressed blood vessel ingrowth into the granulation tissue, whereas holes completely cleared of chitosan amplified angiogenesis vs microdrill-only (P = 0.049), an effect enhanced by IIa. Chitosan implant suppressed strong Col-I, Col-II, and GAG accumulation that occurred spontaneously in drill-only bone defects (P < 0.005) and coagulation factors did not alter this effect. CONCLUSIONS: Subchondral angiogenesis is promoted by chitosan implant clearance. Chitosan implant treatment suppresses fibrocartilage scar tissue formation, and promotes bone remodeling, which allows more blood vessel migration and woven bone repair towards the cartilage lesion area.

Bone marrow stimulation induces greater chondrogenesis in trochlear vs condylar cartilage defects in skeletally mature rabbits.

OBJECTIVE: The aim of this study was to compare the early repair response of cartilage defects in trochlea (TR) and medial femoral condyle (MFC) at 2-3 weeks after bone marrow stimulation. DESIGN: Bilateral full-thickness cartilage defects were generated in central trochlear groove and MFC of skeletally mature rabbits. Four subchondral perforations were made on each defect, either by microfracture to 2 mm deep, or by drilling to 2 mm or 6 mm deep. Rabbits were sacrificed either on Day 14 post-operatively or on Day 21. Defects were analyzed by histology, stereology, histomorphometry and micro-computed tomography (CT). Intact femurs (N = 4) served as controls. RESULTS: Stromal cell density recruitment was similar in all defects, irrespective of defect location and surgical techniques used. There was a robust appearance of chondrocytes at Day 21 in TR defects with significantly higher volume fraction of chondrocytes in TR compared to MFC (P = 0.013). Chondrogenic foci were observed in marrow penetrating holes, with a significantly higher frequency and larger foci in TR vs MFC defects at Day 21 (P = 0.043 and P = 0.0014, respectively). Micro-CT analysis showed that deep drilling elicited significantly more mineralized bone fill compared to shallower perforations at 2 and 3 weeks repair (all at P ≤ 0.0008). CONCLUSIONS: Bone marrow stimulation induced greater chondrogenesis in TR vs MFC defects in adult rabbits, with more chondrocytes and larger chondrogenic foci appearing in TR vs MFC on Day 21 post-operation.

Temporal and spatial modulation of chondrogenic foci in subchondral microdrill holes by chitosan-glycerol phosphate/blood implants.

OBJECTIVE: Subchondral drilling initiates a cartilage repair response involving formation of chondrogenic foci in the subchondral compartment. The purpose of this study was to structurally characterize these sites of chondrogenesis and to investigate the effects of chitosan-glycerol phosphate (GP)/blood implants on their formation. METHOD: Thirty-two New Zealand White rabbits received bilateral cartilage defects bearing four subchondral drill holes. One knee per rabbit was treated by solidifying a chitosan-GP/blood implant over the defect. After 1-56 days of repair, chondrogenic foci were characterized by histostaining and immunostaining. Collagen fiber orientation was characterized by polarized light microscopy. RESULTS: Glycosaminoglycan and collagen type II were present throughout the foci while the upper zone expressed collagen type I and the lower zone collagen type X. Large chondrogenic foci had a stratified structure with flatter cells closer to the articular surface, and round or hypertrophic chondrocytes deeper in the drill holes that showed signs of calcification after 3 weeks of repair in control defects. Markers for pre-hypertrophic chondrocytes (Patched) and for proliferation (Ki-67) were detected within foci. Some cells displayed a columnar arrangement where collagen was vertically oriented. For treated defects, chondrogenic foci appeared 1-3 weeks later, foci were nascent and mature rather than resorbing, and foci developed closer to the articular surface. CONCLUSIONS: Chondrogenic foci bear some similarities to growth cartilage and can give rise to a repair tissue that has similar zonal stratification as articular cartilage. The temporal and spatial formation of chondrogenic foci can be modulated by cartilage repair therapies.

Structural characteristics of the collagen network in human normal, degraded and repair articular cartilages observed in polarized light and scanning electron microscopies.

OBJECTIVE: This study characterizes collagen organization (CO) in human normal (n = 6), degraded (n = 6) and repair (n = 22) cartilages, using polarized light (PLM) and scanning electron (SEM) microscopies. DESIGN: CO was assessed using a recently developed PLM-CO score (Changoor et al. Osteoarthritis Cartilage 2011;19:126-35), and zonal proportions measured. SEM images were captured from locations matched to PLM. Fibre orientations were assessed in SEM and compared to those observed in PLM. CO was also assessed in individual SEM images and combined to generate a SEM-CO score for overall CO analogous to PLM-CO. Fibre diameters were measured in SEM. RESULTS: PLM-CO and SEM-CO scores were correlated, r = 0.786 (P < 0.00001, n = 32), after excluding two outliers. Orientation observed in PLM was validated by SEM since PLM/SEM correspondence occurred in 91.6% of samples. Proportions of the deep (DZ), transitional (TZ) and superficial (SZ) zones averaged 74.0 ± 9.1%, 18.6 ± 7.0%, and 7.3 ± 1.2% in normal, and 45.6 ± 10.7%, 47.2 ± 10.1% and 9.5 ± 3.4% in degraded cartilage, respectively. Fibre diameters in normal cartilage increased with depth from the articular surface [55.8 ± 9.4 nm (SZ), 87.5 ± 1.8 nm (TZ) and 108.2 ± 1.8 nm (DZ)]. Fibre diameters were smaller in repair biopsies [60.4 ± 0.7 nm (SZ), 63.2 ± 0.6 nm (TZ) and 67.2 ± 0.8 nm (DZ)]. Degraded cartilage had wider fibre diameter ranges and bimodal distributions, possibly reflecting new collagen synthesis and remodelling or collagen fibre unravelling. Repair tissues revealed the potential of microfracture-based repair procedures to produce zonal CO resembling native articular cartilage structure. Values are reported as mean ± 95% confidence interval. CONCLUSION: This detailed assessment of collagen architecture could benefit the development of cartilage repair strategies intended to recreate functional collagen architecture.

Screening BRCA1 and BRCA2 unclassified variants for splicing mutations using reverse transcription PCR on patient RNA and an ex vivo assay based on a splicing reporter minigene.

BACKGROUND: Many unclassified variants (UV) of BRCA1 or BRCA2 may have an effect on pre-mRNA splicing. Patient blood samples suitable for RNA extraction are not always available for testing UVs at the RNA level. METHODS: Analyses of RNA from patient peripheral blood were performed, using a one-step reverse transcriptase-PCR (RT-PCR) protocol, and were compared with an ex vivo splicing assay based on PCR-amplified patient DNA inserted into a splicing reporter minigene. Using both methods 20 variants found in 17 patients were examined. RESULTS: Data from patient RNA and from the minigene assay were fully concordant, but the ex vivo splicing assay, which is monoallelic, clarified several ambiguities in the patient RNA data. Two intronic variants induced strong splicing defects: BRCA1 c.4987-5T-->A (IVS16-5T-->A) induced exon 17 skipping and BRCA2 c.316+5G-->C (IVS3+5G-->C) induced complete skipping of exon 3. Of the exonic variants, BRCA2 c.7805G-->C (p.Arg2602Thr), at the last base of exon 16, induced both exon skipping and activation of a cryptic exonic donor site, and BRCA2 c.8023A-->G (p.Ile2675Val) generated a strong donor site within exon 18. These four variants were thus classified as pathogenic, because of the total absence of a normal transcript from the corresponding allele. Variant BRCA2 c.9501+3A-->T (IVS25+3A-->T) induced incomplete skipping of exon 25, suggesting a mutation with incomplete penetrance, and BRCA2 c.8257_8259del (p.Leu2753del) modified the alternative splicing of exons 17 and 18. CONCLUSIONS: We show that functional analysis using a splicing reporter minigene is sensitive and specific, and should be used for initial screening of potential splicing defects, especially when patient RNA is not readily available.

[Routine use of serial plasmatic CA 15-3 determinations during the follow-up of patients treated for breast cancer. Evaluation as factor of early diagnosis of recurrence]. / Utilisation en routine du CA 15.3 dans la surveillance des patientes traitées pour un cancer du sein invasif. Impact en termes de diagnostic précoce de récidive.

PURPOSE: at our institution, CA 15-3 assays are routinely used for the early diagnosis of recurrence during the follow-up of patients treated for breast cancer, although published guidelines do not recommend this procedure. So, we decided to totally re-assess the usefulness of this policy. PATIENTS AND METHODS: all records of patients presenting a first recurrence, local (50 cases) or metastatic (88 cases), of breast cancer during 2003 were re-examined. An increase in CA 15-3 concentration of more than 25% was considered significant. RESULTS: an increase was observed in 18% of non metastatic recurrences. These increases had a prognostic value. CA 15-3 levels remained stable in 23% of metastasis cases and increased in 77%. In 14% of cases, the increase in CA 15-3 levels confirmed a clinically or radiologically suspected metastasis. Moreover, increased CA 15-3 levels in the absence of suggestive clinical or radiological signs led to the diagnosis in 18% of metastasis, 50% of which involved the bone. CONCLUSION: our study demonstrates that CA 15-3 is useful for the early diagnosis of recurrence. Eighteen per cent of metastases were diagnosed by a marker increase alone. CA 15-3 assays could be useful in the early management of these metastases in patients treated for breast cancer.

Injectable chitosan-platelet-rich plasma implants to promote tissue regeneration: in vitro properties, in vivo residence, degradation, cell recruitment and vascularization.

The purpose of this study was to develop freeze-dried chitosan formulations that can be solubilized in platelet-rich plasma (PRP) to form injectable implants for tissue repair. A systematic approach to adjust formulation parameters, including chitosan number average molar mass (Mn ), chitosan concentration and lyoprotectant concentration, was undertaken to identify compositions that would rapidly (< 1 min) and completely solubilize in PRP, would have paste-like handling properties upon solubilization and coagulate rapidly (< 5 min) to form solid chitosan-PRP hybrid implants that are stable and homogenous. Freeze-dried cakes containing calcium chloride, as well as distinct chitosan Mn , chitosan concentration and lyoprotectant concentration, were prepared. PRP was used to solubilize the freeze-dried cakes and assess in vitro and in vivo performance, the latter as dorsal subcutaneous injections into New Zealand White rabbits. Freeze-dried polymer formulations containing low and medium chitosan Mn and concentrations were rapidly and completely solubilized in PRP. The paste-like chitosan-PRP mixtures coagulated quickly to form solid chitosan-PRP hybrids, which retracted much less than PRP-only controls. Homogeneous dispersion of chitosan within the hybrid clots was strongly dependent on chitosan Mn , and occurred only with medium Mn chitosan. Chitosan-PRP hybrid clots were resident subcutaneously in vivo until at least 2 weeks while PRP controls were quickly degraded in one day. Compared to PRP alone, chitosan-PRP hybrids had much greater capacity to induce local cell recruitment accompanied by angiogenesis, suggesting a strong potential for their use in regenerative medicine. Copyright © 2017 John Wiley & Sons, Ltd.

Rab3a controls exocytosis in cholecystokinin-secreting cells.

The expression of rab3A and rab3D isoforms in the enteroendocrine, cholecystokinin-secreting, cell lines STC-1 and GLUTag is here demonstrated. In contrast, rab3B is undetectable in these two cell lines, and rab3C is only slightly expressed in GLUTag cells. Using a transient co-transfection system with human growth hormone as reporter protein, we show that overexpression of the GTPase-deficient mutant rab3AQ81L, but not rab3DQ81L, significantly decreases human growth hormone secretory responses to various agonists in STC-1 cells. These results indicate that endocrine cell lines of intestinal origin express rab3A and rab3D proteins, but the GTP-bound form of rab3A only acts as a negative modulator in the control of cholecystokinin secretion from STC-1 cells.

Tetracosactrin vs. methylprednisolone in the prevention of emesis in patients receiving FEC regimen for breast cancer.

0.5 mg tetracosactrin is considered to be equivalent to 40 mg methylprednisolone with regard to the induced cortisol secretion. 97 female breast cancer patients who received their first two FEC courses (epirubicin 50-75 mg/m2, 5-fluorouracil 500 mg/m2, cyclophosphamide 500 mg/m2) entered this randomised crossover study (76 had previously received an adjuvant treatment); tetracosactrin was administered intramuscularly and methylprednisolone intravenously immediately before chemotherapy administration. The tolerability was evaluated using a diary card during 5 days and patients were asked for their preference at the end of the two cycles. There was no difference either for vomiting (dry heaves were included) or nausea between the two treatments (the analysis was performed on day 1, the worse day of days 2 and 3 and the worse day of days 4 and 5). At day 1, 49% of the patients experienced no or mild nausea after tetracosactrin and 62% after methylprednisolone (not significant) (first period analysis); a complete control of vomiting (including dry heaves) was observed in 49% of the patients after tetracosactrin and 53% after methylprednisolone (not significant). No difference was observed between patients with or without previous chemotherapy. However, slightly more patients preferred tetracosactrin (P = 0.048).

Expression of opioid peptides in cells and stroma of human breast cancer and adenofibromas.

The expression of beta-endorphin, Met-enkephalin and Leu-enkephalin was studied in 63 malignant and benign human breast tumors using immunohistochemical methods. Among invasive ductal carcinomas, 93% were positive for beta-endorphin, 87% for Leu-enkephalin and 90% for Met-enkephalin, in both the tumor stroma and the cell bodies. Enkephalin was predominant in cells, whereas endorphin was predominant in stroma. Nearly the same distribution was found in adenofibromas. In pericancerous normal tissue, neuropeptides were predominantly expressed in the stroma. Although the neuropeptide expression is not cancer-specific, it could be cancer-related, since the results suggest that the neuropeptide expression could reflect the host response to cancer cells and not only the cancer cell activity. The possibility of a direct action of the nervous system on stroma reaction and then on cancer cells is discussed.

Galanin inhibits glucagon-like peptide-1 secretion through pertussis toxin-sensitive G protein and ATP-dependent potassium channels in rat ileal L-cells.

The neuropeptide galanin is widely distributed in the gastrointestinal tract and exerts several inhibitory effects, especially on intestinal motility and on insulin release from pancreatic beta-cells. The presence of galanin fibres not only in the myenteric and submucosal plexus but also in the mucosa, prompted us to investigate the regulatory role of galanin, and its mechanism of action, on the secretion of the insulinotropic hormone glucagon-like peptide-1 (GLP-1). Rat ileal cells were dispersed through mechanical vibration followed by moderate exposure to hyaluronidase, DNase I and EDTA, and enriched for L-cells by counterflow elutriation. A 6- to 7-fold enrichment in GLP-1 cell content was registered after elutriation, as compared with the crude cell preparation (929 +/- 81 vs 138 +/- 14 fmol/10(6) cells). L-cells then accounted for 4-5% of the total cell population. Bombesin induced a time-(15-240 min) and dose- (0.1 nM-1 microM) dependent release of GLP-1. Glucose-dependent insulinotropic peptide (GIP, 100 nM), forskolin (10 microM) and the phorbol ester 12-0-tetradecanoylphorbol-13-acetate (TPA, 1 microM) each stimulated GLP-1 secretion over a 1-h incubation period. Galanin (0.01-100 nM) induced a dose-dependent inhibition of bombesin- and of GIP-stimulated GLP-1 release (mean inhibition of 90% with 100 nM galanin). Galanin also dose-dependently inhibited forskolin-induced GLP-1 secretion (74% of inhibition with 100 nM galanin), but not TPA-stimulated hormone release. Pretreatment of cells with 200 ng/ml pertussis toxin for 3 h, or incubation with the ATP-sensitive K+ channel blocker disopyramide (200 microM), prevented the inhibition by galanin of bombesin- and GIP-stimulated GLP-1 secretion. These studies indicate that intestinal secretion of GLP-1 is negatively controlled by galanin, that acts through receptors coupled to pertussis toxin-sensitive G protein and involves ATP-dependent K+ channels.

Inhibition of active E rosette forming T lymphocytes by hyaluronic acid. Evidence of a receptor for hyaluronic acid on a lymphocyte subpopulation.

Previous studies have shown the inhibition of active E rosette forming T lymphocytes by a mesenchyme associated antigen. Recent results clearly indicated that this antigen consisted in the association of a glycoprotein named hyaluronectin with hyaluronic acid. Using the active E rosette technique of Wybran and Fudenberg, we have studied the action of hyaluronic acid on T lymphocytes. We obtained evidence of the partial inhibition of active E rosette formation by hyaluronic acid in 19 of 25 healthy subjects. Among them, inhibition percentage was 45 +/- 4. This inhibition remained significant at a concentration of 22.5 micrograms/ml hyaluronic acid. Hyaluronic acid was labelled with peroxidase by the glutaraldehyde technique: 17 +/- 7% lymphocytes were stained by this preparation. The preincubation of peroxydase labelled hyaluronic acid by brain hyaluronectin lowered this staining. This is in agreement with the presence of hyaluronectin on a subpopulation of lymphocytes as it was shown by immunofluorescence techniques. In conclusion, a receptor for hyaluronic acid (hyaluronectin) was detected on a proportion of lymphocytes. These results suggest that hyaluronic acid could have an immunosuppressive activity.

Neuroendocrine primary small cell carcinoma of the breast. Report of a case and review of the literature.

One case of breast neuroendocrine primary small cell carcinoma with light microscopic and immunohistochemical findings is reported. The patient died of unrelated disease 21 months after diagnosis and treatment by modified radical mastectomy, radiotherapy and subsequent chemotherapy. Immunohistochemical studies revealed cytokeratin and neuroendocrine markers (chromogranin, neuron-specific enolase) immunostaining on tumoral cells. Expression for neuropeptides (met-enkephalin, leu-enkephalin, beta-endorphin) and CALLA antigen was found. Based on this case report and six other previously reported cases, breast neuroendocrine primary small cell carcinoma appears to be a very aggressive tumor for which no firm conclusions regarding treatment can be drawn.

Role of equalisation mammography of dense breasts.

Parenchymal patterns characteristic of dense breasts are known to degrade the mammographic detection of small breast cancers and microcalcifications. This arises from large variations in exposure of the film, resulting in reduced image contrast over areas of suboptimal exposure. Based on sensitometric measurements of mammograms from a typical patient population, it is shown that over 60% of a typical mammogram in Wolfe's DY classification was found to be exposed suboptimally, suggesting a significant margin for improving mammography for these patients. In order to address this problem, a prototype mammographic version of scanning equalisation radiography (MSER) has been developed, which delivers a patient-specific spatially non-uniform distribution of breast exposure, adjusted to maintain optimal film exposure and contrast over the entire mammogram. Anthropomorphic phantom MSER images show a marked improvement in subjective image quality relative to conventional mammograms, while exhibiting a similar radiation risk. The detection of small microcalcifications and fibrils over clinically significant breast densities is found to be improved by factors eight and four, respectively. Such a system may be clinically practical through the use of multiple-beam equalisation methods with available X-ray tube technology.

[Li-Fraumeni syndrome: update, new data and guidelines for clinical management]. / Le syndrome de Li-Fraumeni: mise au point, données nouvelles et recommandations pour la prise en charge.

The Li-Fraumeni syndrome (LFS) is an inherited form of cancer, affecting children and young adults, and characterized by a wide spectrum of tumors, including soft-tissue and bone sarcomas, brain tumours, adenocortical tumours and premenopausal breast cancers. In most of the families, LFS results from germline mutations of the tumor suppressor TP53 gene encoding a transcriptional factor able to regulate cell cycle and apoptosis when DNA damage occurs. Recently, germline mutations of hCHK2 encoding a kinase, regulating cell cycle via Cdc25C and TP53, were identified in affected families. The LFS working group recommendations are the following: (i) positive testing (screening for a germline TP53 mutation in a patient with a tumor) can be offered both to children and adults in the context of genetic counseling associated to psychological support, to confirm the diagnosis of LFS on a molecular basis. This will allow to offer to the patient a regular clinical review in order to avoid a delay to the diagnosis of another tumor; (ii) the 3 indications for positive testing are: a proband with a tumor belonging to the narrow LFS spectrum and developed before age 36 and, at least, first- or second-degree relative with a LFS spectrum tumor, before age 46, or a patient with multiple primary tumors, 2 of which belonging to the narrow LFS spectrum, the first being developed before 36 or a child with an adenocortical tumour; (iii) presymptomatic testing must be restricted to adults; (iv) the young age of onset of the LFS tumors the prognosis of some tumors, the impossibility to ensure an efficient early detection and the risk for mutation carriers to develop multiple primary tumors justify that prenatal diagnosis might be considered in affected families.

[Some insights on the history of children's drawings]. / Quelques aperçus sur l'histoire du dessin d'enfant.

Both children's use of drawings and their interpretation in psychopathology have a history and even a pre-history. The author comments on the few drawn documents he could gather concerning the latter. He examines the birth of children drawings in psychology, psychiatry and education at the end of the XIXth century and the turn of the XXth. He finally recalls the first traces of its use by psychoanalysis, mainly in France. Some excursi are joined relative to the conditions of discovery of pre-historic art, to the relationship between modern art and child "primitivism", and to the themes of child drawings within cartoons.

Acute Osteoclast Activity following Subchondral Drilling Is Promoted by Chitosan and Associated with Improved Cartilage Repair Tissue Integration.

OBJECTIVE: Cartilage-bone integration is an important functional end point of cartilage repair therapy, but little is known about how to promote integration. We tested the hypothesis that chitosan-stabilized blood clot implant elicits osteoclasts to drilled cartilage defects and promotes repair and cartilage-bone integration. DESIGN: Bilateral trochlear defects in 15 skeletally mature rabbit knees were microdrilled and then treated with chitosan-glycerol phosphate (GP)/blood implant with fluorescent chitosan tracer and thrombin to accelerate in situ solidification or with thrombin alone. Chitosan clearance, osteoclast density, and osteochondral repair were evaluated at 1, 2, and 8 weeks at the outside, edge, and through the proximal microdrill holes. RESULTS: Chitosan was retained at the top of the drill holes at 1 week as extracellular particles became internalized by granulation tissue cells at 2 weeks and was completely cleared by 8 weeks. Osteoclasts burst-accumulated at microdrill hole edges at 1 week, in new woven bone at the base of the drill holes at 2 weeks, and below endochondral cartilage repair at 8 weeks. Implants elicited 2-fold more osteoclasts relative to controls (P < 0.001), a more complete drill hole bone repair, and improved cartilage-bone integration and histological tissue quality. Treated and control 8-week cartilage repair tissues contained 85% collagen type II. After 8 weeks of repair, subchondral osteoclast density correlated positively with bone-cartilage repair tissue integration (P < 0.0005). CONCLUSIONS: Chitosan-GP/blood implant amplified the acute influx of subchondral osteoclasts through indirect mechanisms, leading to significantly improved repair and cartilage-bone integration without inducing net bone resorption. Osteoclasts are cellular mediators of marrow-derived cartilage repair integration.