Procedures
effusion
EFFUSION
The cellular harvesting, final product preparation, injection procedures and CSI were standardized across the five study sites. This was done through training courses before study initiation as well as subsequent monitoring by the lead site. Of note, the BMAC and SVF were fresh autologous products while the UCT were cryopreserved, purified MSCs manufactured from donated allogeneic umbilical cord tissue in a cGMP (current Good Manufacturing Practice) facility. Supplementary Material All procedures were done in an outpatient clinic setting with another clean room used for point-of-care laboratory testing. No cGMP facility was used for the actual treatment portion of this study. The subject was positioned supine on an examination table with foam roller/bolster under knee. The injectate was prepared by the research team with opaque tape wrapped around the 10-ml syringe to maintain the blinding. If an effusion was present based on clinical and ultrasound examination, 2 ml of ropivicaine 0.2% was injected between the skin and down to the joint capsule. Following this, an 18-gauge needle was used to aspirate any joint fluid via a superior lateral approach under direct ultrasound guidance. Following joint aspiration (if performed), the solution of either corticosteroids, BMAC, SVF or UCT was injected under ultrasound guidance. Following the injection, the knee was passively moved from extension to flexion three to four times to help spread the injectate and patients were instructed to remain supine for 10 min.All final cellular products were tested in the clinic for total nucleated cell count, cell viability and endotoxin levels to determine if release criteria were met before injection. These tests were performed using a Nexcelom Auto 2000 device for cell counting and cell viability and the Charles River Endosafe Nexgen PTS for endotoxin testing. A 1-ml aliquot of each cellular product was separated from the final injection preparation for the release criteria testing and FDA requirements. If the final product did not meet the required release criteria depicted in Table Cellular diversity in BMAC and SVF samples and cellular heterogeneity in UCT-MSC samples were evaluated by single-cell RNA sequencing. The single-cell RNA sequencing libraries were prepared with 10x Genomics Chromium platform using 3′ V3.1 kit. The sequencing was done on Illumina Novaseq 6000 platform with an S4 kit. We used a modified SEURAT pipeline to analyze the samples