The synovial fluid from patients with focal cartilage defects contains mesenchymal stem/stromal cells and macrophages with pro- and anti-inflammatory phenotypes

Objective The synovial fluid (SF) of patients with focal cartilage defects contains a population of poorly characterised cells that could have pathophysiological implications in early osteoarthritis and joint tissue repair. We have examined the cells within SF of such joints by determining their chondrogenic capacity following culture expansion and establishing the phenotypes of the macrophage subsets in non-cultured cells. Design Knee SF cells were obtained from 21 patients receiving cell therapy to treat a focal cartilage defect. Cell surface immunoprofiling for stem cell and putative chondrogenic markers, and the expression analysis of key chondrogenic and hypertrophic genes were conducted on culture-expanded SF cells prior to chondrogenesis. Flow cytometry was also used to determine the macrophage subsets in freshly isolated SF cells. Results Immunoprofiling revealed positivity for the monocyte/macrophage marker (CD14), the haematopoietic/endothelial cell marker (CD34) and mesenchymal stem/stromal cell markers (CD73, CD90, CD105) on culture expanded cells. We found strong correlations between the presence of CD14 and the vascular cell adhesion marker, CD106 (r=0.81, p=0.003). Collagen type II expression after culture expansion positively correlated with GAG production (r=0.73, p=0.006), whereas CD90 (r=-0.6, p=0.03) and CD105 (r=-0.55, p=0.04) immunopositivity were inversely related to GAG production. Freshly isolated SF cells were positive for both pro- (CD86) and anti-inflammatory markers (CD163 and CD206). Conclusions The cellular content of the SF from patients with focal cartilage injuries is comprised of a heterogeneous population of reparative and inflammatory cells. Additional investigations are needed to understand the role played by these cells in the attempted repair and inflammatory process in diseased joints

Autologous chondrocyte implantation- derived synovial fluids display distinct responder and non-responder proteomic profiles

Background Autologous chondrocyte implantation (ACI) can be used in the treatment of focal cartilage injuries to prevent the onset of osteoarthritis (OA). However, we are yet to understand fully why some individuals do not respond well to this intervention. Identification of a reliable and accurate biomarker panel that can predict which patients are likely to respond well to ACI is needed in order to assign the patient to the most appropriate therapy. This study aimed to compare the baseline and mid-treatment proteomic profiles of synovial fluids (SFs) obtained from responders and non-responders to ACI. Methods SFs were derived from 14 ACI responders (mean Lysholm improvement of 33 (17–54)) and 13 non-responders (mean Lysholm decrease of 14 (4–46)) at the two stages of surgery (cartilage harvest and chondrocyte implantation). Label-free proteome profiling of dynamically compressed SFs was used to identify predictive markers of ACI success or failure and to investigate the biological pathways involved in the clinical response to ACI. Results Only 1 protein displayed a =2.0-fold differential abundance in the preclinical SF of ACI responders versus non-responders. However, there is a marked difference between these two groups with regard to their proteome shift in response to cartilage harvest, with 24 and 92 proteins showing =2.0-fold differential abundance between Stages I and II in responders and non-responders, respectively. Proteomic data has been uploaded to ProteomeXchange (identifier: PXD005220). We have validated two biologically relevant protein changes associated with this response, demonstrating that matrix metalloproteinase 1 was prominently elevated and S100 calcium binding protein A13 was reduced in response to cartilage harvest in non-responders. Conclusions The differential proteomic response to cartilage harvest noted in responders versus non-responders is completely novel. Our analyses suggest several pathways which appear to be altered in non-responders that are worthy of further investigation to elucidate the mechanisms of ACI failure. These protein changes highlight many putative biomarkers that may have potential for prediction of ACI treatment success

Two independent proteomic approaches provide a comprehensive analysis of the synovial fluid proteome response to Autologous Chondrocyte Implantation

Background: Autologous chondrocyte implantation (ACI) has a failure rate of approximately 20%, but it is yet to be fully understood why. Biomarkers are needed that can pre-operatively predict in which patients it is likely to fail, so that alternative or individualised therapies can be offered. We previously used label-free quantitation (LF) with a dynamic range compression proteomic approach to assess the synovial fluid (SF) of ACI responders and non-responders. However, we were able to identify only a few differentially abundant proteins at baseline. In the present study, we built upon these previous findings by assessing higher-abundance proteins within this SF, providing a more global proteomic analysis on the basis of which more of the biology underlying ACI success or failure can be understood. Methods: Isobaric tagging for relative and absolute quantitation (iTRAQ) proteomic analysis was used to assess SF from ACI responders (mean Lysholm improvement of 33; n = 14) and non-responders (mean Lysholm decrease of 14; n = 13) at the two stages of surgery (cartilage harvest and chondrocyte implantation). Differentially abundant proteins in iTRAQ and combined iTRAQ and LF datasets were investigated using pathway and network analyses. Results: iTRAQ proteomic analysis confirmed our previous finding that there is a marked proteomic shift in response to cartilage harvest (70 and 54 proteins demonstrating ≥ 2.0-fold change and p < 0.05 between stages I and II in responders and non-responders, respectively). Further, it highlighted 28 proteins that were differentially abundant between responders and non-responders to ACI, which were not found in the LF study, 16 of which were altered at baseline. The differential expression of two proteins (complement C1s subcomponent and matrix metalloproteinase 3) was confirmed biochemically. Combination of the iTRAQ and LF proteomic datasets generated in-depth SF proteome information that was used to generate interactome networks representing ACI success or failure. Functional pathways that are dysregulated in ACI non-responders were identified, including acute-phase response signalling. Conclusions: Several candidate biomarkers for baseline prediction of ACI outcome were identified. A holistic overview of the SF proteome in responders and non-responders to ACI  has been profiled, providing a better understanding of the biological pathways underlying clinical outcome, particularly the differential response to cartilage harvest in non-responders

Global patient outcomes after elective surgery: prospective cohort study in 27 low-, middle- and high-income countries.

BACKGROUND: As global initiatives increase patient access to surgical treatments, there remains a need to understand the adverse effects of surgery and define appropriate levels of perioperative care. METHODS: We designed a prospective international 7-day cohort study of outcomes following elective adult inpatient surgery in 27 countries. The primary outcome was in-hospital complications. Secondary outcomes were death following a complication (failure to rescue) and death in hospital. Process measures were admission to critical care immediately after surgery or to treat a complication and duration of hospital stay. A single definition of critical care was used for all countries. RESULTS: A total of 474 hospitals in 19 high-, 7 middle- and 1 low-income country were included in the primary analysis. Data included 44 814 patients with a median hospital stay of 4 (range 2-7) days. A total of 7508 patients (16.8%) developed one or more postoperative complication and 207 died (0.5%). The overall mortality among patients who developed complications was 2.8%. Mortality following complications ranged from 2.4% for pulmonary embolism to 43.9% for cardiac arrest. A total of 4360 (9.7%) patients were admitted to a critical care unit as routine immediately after surgery, of whom 2198 (50.4%) developed a complication, with 105 (2.4%) deaths. A total of 1233 patients (16.4%) were admitted to a critical care unit to treat complications, with 119 (9.7%) deaths. Despite lower baseline risk, outcomes were similar in low- and middle-income compared with high-income countries. CONCLUSIONS: Poor patient outcomes are common after inpatient surgery. Global initiatives to increase access to surgical treatments should also address the need for safe perioperative care. STUDY REGISTRATION: ISRCTN5181700

Improving care for people with heart failure in Uganda: serial in-depth interviews with patients' and their health care professionals

Abstract Background The short prognosis of patients with advanced heart failure (HF) and the associated multidimensional distress as illustrated in literature from high income countries necessitates the integration of palliative care into the care of advanced HF patients to address these needs and improve their quality of life. These needs, which are subjective, have not been described from the patients’ and health care professionals’(HPs) view point in the Ugandan setting, a low income country with a different socio-cultural context. This study aimed at bridging this gap in knowledge by eliciting patients’ and HPs’ views of HF patients’ needs over the course of their illness to enable generalists, cardiologists and palliative care clinicians to develop guidelines to provide patient-centred realistic care in Uganda. Methods Serial qualitative in-depth interviews were conducted with HF patients who were purposively sampled and recruited in Mulago National Referral Hospital (MNRH) until thematic saturation. In-depth interviews were conducted at three time points with intervals of 3 month between interviews over the course of their illness in the hospital and their home context. One-off interviews were conducted with HPs that manage HF in MNRH. We used a grounded theory approach in data analysis. The Uganda National Council of science and technology approved the research. Results Forty-eight interviews were conducted with 21 patients and their carers and eight interviews with their HPs. Multidimensional needs including physical, psychological, social, spiritual and information needs were identified. These highlighted the underpinning need to have normal functioning, control, to cope and adapt to a changed life and to find meaning. Spiritual needs were less recognised by HPs than the other multidimensional needs. Information needs were commonly unmet. Patients and HPs suggested improvements in care that were congruent with the recommendations in chronic disease care and the six pillars of the WHO health systems strengthening approach. Conclusion Management of HF in Uganda requires an approach that targets multidimensional needs, embraces multidisciplinary care and strengthens health systems which are all important tenets of palliative care

Hulme PROTEOMIC IDENTIFICATION OF PREDICTIVE AUTOLOGOUS CHONDROCYTE IMPLANTATION BIOMARKERS IN PLASMA

Stratification is required to ensure that only those patients likely to benefit, receive Autologous Chondrocyte Implantation (ACI); ideally by assessing a biomarker in the blood. This study aimed to assess differences in the plasma proteome of individuals who respond well or poorly to ACI. Isobaric tag for relative and absolute quantitation (ITRAQ) mass spectrometry and label-free proteomics analyses were performed in tandem as described previously by our group (Hulme et al., 2017; 2018; 2021) using plasma collected from ACI responders (n=10) compared with non-responders (n=10) at each stage of surgery (Stage I, cartilage harvest and Stage II, cell implantation). iTRAQ using pooled plasma detected 16 proteins that were differentially abundant at baseline in ACI responders compared with non-responders (n=10) (≥±2.0 fold; p<0.05). Responders demonstrated a mean Lysholm (patient reported functional score from 0–100) improvement of 33±13 and non-responders a mean worsening of −13±13 points. The most pronounced plasma proteome shift was seen in response to Stage I surgery in ACI non-responders, with 48 proteins being differentially abundant between the two surgical procedures. We have previously noted this marked shift in response to initial surgery in the SF of ACI non-responders, several of these proteins were associated with the Acute Phase Response. One of these proteins, clusterin, could be confirmed in patients’ plasma using an independent immunoassay using individual samples. Label-free proteomic data from individual samples identified only cartilage acidic protein-1 (known to associate with osteoarthritis progression) to be significantly more abundant at Stage I in the plasma of non-responders. This study indicates that proteins can be identified within the plasma that have potential use in ACI patient stratification. Further work is required to validate the findings of this discovery-phase work in larger ACI cohorts