Light as a drug: prospective randomized evaluation and comparison of the effect of decreased illumination on visual recovery after cataract surgery.

PURPOSE: To compare the effect of decreased illumination on the rate of postoperative visual recovery, and the incidence of cystoid macular edema (CME) with surgical visualization achieved with a traditional analog operating microscope compared with a 3D digital visualization system. SETTING: Ambulatory surgery center, New York. DESIGN: Prospective, randomized, consecutive, single-surgeon series. METHODS: Patients undergoing routine cataract surgery were randomized into either (1) visualization through the binoculars of a standard operating microscope ("traditional group") or (2) visualization through a 3D digital visualization system affixed to the same operating microscope ("digital group"). Note was made in each case of light intensity used, light exposure time, cumulative dissipated energy (CDE), femtosecond laser use, preoperative medical and ocular conditions, intraoperative and/or postoperative complications, and preoperative and postoperative visual acuities and optical coherence tomography confirmed CME. RESULTS: The study comprised 118 eyes in the traditional group and 96 eyes in the digital group. There were no differences in preoperative visual acuity, light exposure time, CDE, or femtosecond laser use between groups, but the light intensity used in the digital group was significantly less (19.5% ± 0.5%) than in the traditional group (48.6% ± 0.6%; P < .001). Furthermore, the digital group achieved a better decimal postoperative day 1 visual acuity (0.60 ± 0.03) with less rates of CME (2.1%) when compared with that of the traditional group (0.51 ± 0.02, P = .03; and 9.2%, P = .03), respectively. CONCLUSIONS: Visual recovery and CME rates were significantly better in patients who underwent cataract surgery assisted by the 3D digital visualization platform without an increase in complications or surgical time.

Autologous chondrocyte implantation provides good long-term clinical results in the treatment of knee osteoarthritis: a systematic review.

PURPOSE: To evaluate the mid- and long-term efficacy of autologous chondrocyte implantation (ACI) and matrix-assisted chondrocyte implantation (MACI) to treat patients with knee cartilage defects in the presence of osteoarthritis (OA). METHODS: PubMed and Cochrane databases were systematically searched for studies describing the treatment of knee OA with ACI or MACI (Kellgren-Lawrence (KL) ≥ 1, minimum follow-up 36 months). Results were reported according to the Preferred Reporting Items for Systematic Reviews and Meta-Analysis guidelines and included Lysholm, Western Ontario McMaster University and International Knee Documentation Committee scores. RESULTS: Of the 127 full-text articles assessed for eligibility, only five studies were selected based on inclusion/exclusion criteria (2 on ACI and 3 on MACI). In both groups, the defects were mainly located at femoral level, size 2.2-15.1 cm2 in the ACI and 2.0-7.6 cm2 in the MACI group. ACI was mostly used for patients affected by KL I, whereas MACI for patients with KL II-IV. The data obtained from 235 patients (161 ACI, 74 MACI) showed that ACI and MACI sustained stable clinical improvements up to 11 and 15 years, respectively, with a failure rate of about 10% up to 11 years. Scarce biological details regarding chondrocyte implantation were reported. CONCLUSIONS: ACI and MACI procedures for the treatment of knee cartilage lesions associated to OA showed long-term success and allowed delaying arthroplasty. Additional trials reporting homogenous data and precise patient characterization are needed to conduct an effective literature meta-analysis and identify the clinical relevance of these procedures. LEVEL OF EVIDENCE: IV.

Stability of housekeeping genes in human intervertebral disc, endplate and articular cartilage cells in multiple conditions for reliable transcriptional analysis.

Quantitative gene expression analysis is widely used to evaluate the expression of specific tissue markers. To obtain reliable data it is essential to select stable housekeeping genes whose expression is not influenced by the anatomical origin of cells or by the culture conditions. No studies have evaluated housekeeping gene stability in intervertebral disc (IVD) cells and only few studies using cartilaginous endplate (CEP) and articular cartilage (AC) cells are present in the literature. We analysed the stability of four candidate housekeeping genes (GAPDH, TBP, YWHAZ and RPL13A) in human cells isolated from nucleus pulposus (NP) and annulus fibrosus (AF), CEP and AC. Cell isolation, expansion, cryoconservation, and differentiation in 3D pellets were tested. GeNorm, NormFinder, BestKeeper tools and the comparative ΔCt method were used to evaluate housekeeping gene stability. In each cell population, TBP alone or combined with YWHAZ was identified as the best normaliser in both monolayer and 3D pellets. GAPDH was the best performer only for AC cells in monolayer. In most culture conditions considering groups of two or more cell types, TBP was the most stable and YWHAZ was the second choice. GAPDH was the best performer only in 3D pellets with factors for AC and AF combined with CEP cells. RPL13A was the most stable only for AF with CEP cells at isolation. Our findings will be useful to properly design the experimental set-up of studies involving IVD, CEP or AC cells in different culture conditions, in order to obtain accurate and high quality data from quantitative gene expression analysis.

In Vivo Bone Formation Within Engineered Hydroxyapatite Scaffolds in a Sheep Model.

Large bone defects still represent a major burden in orthopedics, requiring bone-graft implantation to promote the bone repair. Along with autografts that currently represent the gold standard for complicated fracture repair, the bone tissue engineering offers a promising alternative strategy combining bone-graft substitutes with osteoprogenitor cells able to support the bone tissue ingrowth within the implant. Hence, the optimization of cell loading and distribution within osteoconductive scaffolds is mandatory to support a successful bone formation within the scaffold pores. With this purpose, we engineered constructs by seeding and culturing autologous, osteodifferentiated bone marrow mesenchymal stem cells within hydroxyapatite (HA)-based grafts by means of a perfusion bioreactor to enhance the in vivo implant-bone osseointegration in an ovine model. Specifically, we compared the engineered constructs in two different anatomical bone sites, tibia, and femur, compared with cell-free or static cell-loaded scaffolds. After 2 and 4 months, the bone formation and the scaffold osseointegration were assessed by micro-CT and histological analyses. The results demonstrated the capability of the acellular HA-based grafts to determine an implant-bone osseointegration similar to that of statically or dynamically cultured grafts. Our study demonstrated that the tibia is characterized by a lower bone repair capability compared to femur, in which the contribution of transplanted cells is not crucial to enhance the bone-implant osseointegration. Indeed, only in tibia, the dynamic cell-loaded implants performed slightly better than the cell-free or static cell-loaded grafts, indicating that this is a valid approach to sustain the bone deposition and osseointegration in disadvantaged anatomical sites.

Diabetes and the Association of Postoperative Hyperglycemia With Clinical and Economic Outcomes in Cardiac Surgery.

OBJECTIVE: The management of postoperative hyperglycemia is controversial and generally does not take into account pre-existing diabetes. We analyzed clinical and economic outcomes associated with postoperative hyperglycemia in cardiac surgery patients, stratifying by diabetes status. RESEARCH DESIGN AND METHODS: Multicenter cohort study in 4,316 cardiac surgery patients operated on in 2010. Glucose was measured at 6-h intervals for 48 h postoperatively. Outcomes included cost, hospital length of stay (LOS), cardiac and respiratory complications, major infections, and death. Associations between maximum glucose levels and outcomes were assessed with multivariable regression and recycled prediction analyses. RESULTS: In patients without diabetes, increasing glucose levels were associated with a gradual worsening of outcomes. In these patients, hyperglycemia (≥180 mg/dL) was associated with an additional cost of $3,192 (95% CI 1,972 to 4,456), an additional hospital LOS of 0.8 days (0.4 to 1.3), an increase in infections of 1.6% (0.5 to 2.8), and an increase in respiratory complications of 2.6% (0.0 to 5.3). However, among patients with insulin-treated diabetes, optimal outcomes were associated with glucose levels considered to be hyperglycemic (180 to 240 mg/dL). This level of hyperglycemia was associated with cost reductions of $6,225 (-12,886 to -222), hospital LOS reductions of 1.6 days (-3.7 to 0.4), infection reductions of 4.1% (-9.1 to 0.0), and reductions in respiratory complication of 12.5% (-22.4 to -3.0). In patients with non-insulin-treated diabetes, outcomes did not differ significantly when hyperglycemia was present. CONCLUSIONS: Glucose levels <180 mg/dL are associated with better outcomes in most patients, but worse outcomes in patients with diabetes with a history of prior insulin use. These findings support further investigation of a stratified approach to the management of patients with stress-induced postoperative hyperglycemia based on prior diabetes status.

Arthritic and non-arthritic synovial fluids modulate IL10 and IL1RA gene expression in differentially activated primary human monocytes.

OBJECTIVE: Synovitis with an increased presence of macrophages is observed in osteoarthritis (OA) and rheumatoid arthritis (RA). Given the important role of macrophages in arthritis, we investigated the influence of OA and RA synovial fluid (SF) on primary human monocytes (Mo), their lineage precursors. METHOD: Adherent monocytes without any stimulation (Mo(-)) or stimulated with IFN-γ and TNF-α (Mo(IFN-γ/TNF-α)) or IL-4 (Mo(IL-4)) were exposed to SF from 6 donors without any known joint disease (SF-Ctrl), 10 OA donors (SF-OA), and 10 RA donors (SF-RA). The transcriptional expression of IL6, IL1B, TNFA, IL10, CCL18, CD206, and IL1RA was analyzed. RESULTS: Mo(-) exposed to SF-RA had a lower expression of IL10 and a higher expression of IL1RA than when exposed to SF-Ctrl. Mo(IL-4) exposed to SF-RA had a lower expression of IL10 and CCL18 than when exposed to SF-Ctrl and Mo(IFN-γ/TNF-α) were not affected by SF-RA. Mo exposed to SF-OA also expressed less IL10, but only upon stimulation with IL-4, and expressed more IL1RA than when exposed to SF-Ctrl in any condition. CONCLUSION: A lower expression of IL10 may be regarded as a response to less inflammatory conditions since IL10 expression is higher in response to IFN-γ/TNF-α stimulation, probably as a feedback mechanism. Therefore, the lower expression of IL10 and the higher expression of IL1RA in Mo exposed to arthritic than to non-arthritic SF suggest that arthritic SF is mainly reducing the inflammatory responses in Mo. This may mimic the response of monocytes/macrophages recruited to the joint, where feedback mechanisms counteract pro-inflammatory processes.

Donor-matched mesenchymal stem cells from knee infrapatellar and subcutaneous adipose tissue of osteoarthritic donors display differential chondrogenic and osteogenic commitment.

Cell-based therapies have recently been proposed for the treatment of degenerative articular pathologies, such as early osteoarthritis, with an emphasis on autologous mesenchymal stem cells (MSCs), as an alternative to terminally differentiated cells. In this study, we performed a donor-matched comparison between infrapatellar fat pad MSCs (IFP-MSCs) and knee subcutaneous adipose tissue stem cells (ASCs), as appealing candidates for cell-based therapies that are easily accessible during surgery. IFP-MSCs and ASCs were obtained from 25 osteoarthritic patients undergoing total knee replacement and compared for their immunophenotype and differentiative potential. Undifferentiated IFP-MSCs and ASCs displayed the same immunophenotype, typical of MSCs (CD13+/CD29+/CD44+/CD73+/CD90+/CD105+/CD166+/CD31-/CD45-). IFP-MSCs and ASCs showed similar adipogenic potential, though undifferentiated ASCs had higher LEP expression compared to IFP-MSCs (p<0.01). Higher levels of calcified matrix (p<0.05) and alkaline phosphatase (p<0.05) in ASCs highlighted their superior osteogenic commitment compared to IFP-MSCs. Conversely, IFP-MSCs pellets showed greater amounts of glycosaminoglycans (p<0.01) and superior expression of ACAN (p<0.001), SOX9, COMP (p<0.001) and COL2A1 (p<0.05) compared to ASCs pellets, revealing a superior chondrogenic potential. This was also supported by lower COL10A1 (p<0.05) and COL1A1 (p<0.01) expression and lower alkaline phosphatase release (p<0.05) by IFP-MSCs compared to ASCs. The observed dissimilarities between IFP-MSCs and ASCs show that, despite expressing similar surface markers, MSCs deriving from different fat depots in the same surgical site possess specific features. Furthermore, the in vitro peculiar commitment of IFP-MSCs and ASCs from osteoarthritic donors towards the chondrogenic or osteogenic lineage may suggest a preferential use for cartilage and bone cell-based treatments, respectively.

Influence on chondrogenesis of human osteoarthritic chondrocytes in co-culture with donor-matched mesenchymal stem cells from infrapatellar fat pad and subcutaneous adipose tissue.

Co-culture of mesenchymal stem cells (MSCs) and articular chondrocytes (ACs) has been proposed for autologous cartilage cell-based therapies, to overcome the issues associated to limited availability of articular chondrocytes (ACs). To evaluate the potentiality of a co-culture approach in aged osteoarthritic patients, MSCs from infrapatellar fat pad (IFP-MSCs) and knee subcutaneous adipose tissue (ASCs) were co-cultured with donor-matched osteoarthritic, expanded and cryopreserved, ACs in a 75%/25% ratio. Co-cultures were prepared also from nasal chondrocytes (NCs) to evaluate their possible use as an alternative to ACs. Pellets were differentiated for 14 days, using mono-cultures of each cell type as reference. Chondrogenic genes SOX9, COL2A1, ACAN were less expressed in co-cultures compared to ACs and NCs. Total GAGs content in co-cultures did not differ significantly from values predicted as the sum of each cell type contribution corrected for the co-culture ratio, as confirmed by histology. No significant differences were observed for GAGs/DNA in mono-cultures, demonstrating a reduced chondrogenic potential of ACs and NCs. In conclusion, a small percentage of expanded and cryopreserved ACs and NCs did not lead to IFP-MSCs and ASCs chondro-induction. Our results suggest that chondrogenic potential and origin of chondrocytes may play a relevant role in the outcome of co-cultures, indicating a need for further investigations to demonstrate their clinical relevance in the treatment of aged osteoarthritic patients.

Reversine increases multipotent human mesenchymal cells differentiation potential.

Among different human stem cell sources, adult mesenchymal stem cells from bone marrow (BMSCs), and more recently from adipose tissues (ASCs), have shown their capability to differentiate into a variety of different cell types, including osteoblasts, adipocytes, and muscle cells. However, mesenchymal stem cell differentiation toward certain cell types (including skeletal and cardiac muscle), while shown to be achievable, still suffers of low yields and needs to be greatly improved before any therapeutic application could be foreseen. A possible way of achieving this goal is by using a chemical-pharmacological approach to increase stem cell plasticity. Along this line, we envisioned the possibility of pre-treating BMSCs and ASCs with reversine, a synthetic purine that has been shown to induce adult cells de-differentiation. In the current study we tested reversine effects on both BMSCs and ASCs to increase their differentiation toward osteoblasts, smooth and skeletal muscle cells. Reversine pre-treatment, at very low concentration (50 nM), caused a marked increase in the differentiation yields of both BMSCs and ASCs.

Enhanced biological performance of human adipose-derived stem cells cultured on titanium-based biomaterials and silicon carbide sheets for orthopaedic applications.

It is well known that the surface properties of biomaterials may affect bone-healing processes by modulating both cell viability and osteogenic differentiation. In this study we evaluated proliferation and osteogenic differentiation of human adipose-derived stem cells (hASCs) cultured on three prototypes of titanium disks and on thin layers of silicon carbide (SiC-PECVD), a material characterized by a high hardness and wear resistance. Our data indicated that all the tested surfaces supported cell growth, in particular, hASCs seeded on both titanium treated by a double-step etching process (TIT) and titanium modified by two Anodic Spark Deposition processes (TAA) grew better respect to the ones cultured on titanium obtained by KOH alkali etching process on TAA (TAAK). Furthermore, hASCs well colonized SiC-PECVD surface, showing a quite similar viability to cells cultured on plastic (PA). TIT and TAA better supported osteogenic differentiation of hASCs compared to PA, as shown by a marked increase of both alkaline phosphatase activity and calcified extracellular matrix deposition; in contrast TAAK did not positively affect hASCs differentiation. SiC-PECVD did not alter osteogenic differentiation of hASC cells: indeed, ALP and calcium deposition levels were comparable to those of cells cultured on plastic. Furthermore, we observed similar results testing hASCs either pre-differentiated for 14 days in osteogenic medium or directly differentiated on biomaterials. Our study suggests that modifications of titanium surface may improve osteo-integration of implant devices and that SiC-PECVD may represent a valid alternative for the coating of prosthetic devices to reduce wear and metallosis events.

Role of autologous rabbit adipose-derived stem cells in the early phases of the repairing process of critical bone defects.

Adipose-derived stem cells (ASCs) may represent a novel and efficient tool to promote bone regeneration. In this study, rabbit ASCs were expanded in culture and used for the regeneration of full-thickness bone defects in the proximal epiphysis of tibia of 12 New Zealand rabbits. Defects were implanted with graft material as follows: untreated (control), empty hydroxyapatite (HA) disk, ASCs alone, and HA disk seeded with ASCs. Each isolated ASCs population was tested in vitro: they all showed a high proliferation rate, a marked clonogenic ability, and osteogenic differentiation potential. Eight weeks after implantation, macroscopic analyses of all the samples showed satisfactory filling of the lesions without any significant differences in term of stiffness between groups treated with or without cells (p > 0.05). In both the scaffold-treated groups, a good osteointegration was radiographically observed. Even if HA was not completely reabsorbed, ASCs-loaded HA displayed a higher scaffold resorption than the unloaded ones. Histological analyses showed that the osteogenic abilities of the scaffold-treated defects was greater than those of scaffold-free samples, and in particular new formed bone was more mature and more similar to native bone in presence of ASCs. These results demonstrated that autologous ASCs-HA constructs is a potential treatment for the regeneration of bone defects.