Umbilical cord mesenchymal stem cells for COVID-19 acute respiratory distress syndrome: A double-blind, phase 1/2a, randomized controlled trial

Acute respiratory distress syndrome (ARDS) in COVID-19 is associated with high mortality. Mesenchymal stem cells are known to exert immunomodulatory and anti-inflammatory effects and could yield beneficial effects in COVID-19 ARDS. The objective of this study was to determine safety and explore efficacy of umbilical cord mesenchymal stem cell (UC-MSC) infusions in subjects with COVID-19 ARDS. A double-blind, phase 1/2a, randomized, controlled trial was performed. Randomization and stratification by ARDS severity was used to foster balance among groups. All subjects were analyzed under intention to treat design. Twenty-four subjects were randomized 1:1 to either UC-MSC treatment (n = 12) or the control group (n = 12). Subjects in the UC-MSC treatment group received two intravenous infusions (at day 0 and 3) of 100 ± 20 × 106 UC-MSCs; controls received two infusions of vehicle solution. Both groups received best standard of care. Primary endpoint was safety (adverse events [AEs]) within 6 hours; cardiac arrest or death within 24 hours postinfusion). Secondary endpoints included patient survival at 31 days after the first infusion and time to recovery. No difference was observed between groups in infusion-associated AEs. No serious adverse events (SAEs) were observed related to UC-MSC infusions. UC-MSC infusions in COVID-19 ARDS were found to be safe. Inflammatory cytokines were significantly decreased in UC-MSC-treated subjects at day 6. Treatment was associated with significantly improved patient survival (91% vs 42%, P =.015), SAE-free survival (P =.008), and time to recovery (P =.03). UC-MSC infusions are safe and could be beneficial in treating subjects with COVID-19 ARDS

National Institutes of Health–Sponsored Clinical Islet Transplantation Consortium Phase 3 Trial: Manufacture of a Complex Cellular Product at Eight Processing Facilities

Eight manufacturing facilities participating in the National Institutes of Health–sponsored Clinical Islet Transplantation (CIT) Consortium jointly developed and implemented a harmonized process for the manufacture of allogeneic purified human pancreatic islet (PHPI) product evaluated in a phase 3 trial in subjects with type 1 diabetes. Manufacturing was controlled by a common master production batch record, standard operating procedures that included acceptance criteria for deceased donor organ pancreata and critical raw materials, PHPI product specifications, certificate of analysis, and test methods. The process was compliant with Current Good Manufacturing Practices and Current Good Tissue Practices. This report describes the manufacturing process for 75 PHPI clinical lots and summarizes the results, including lot release. The results demonstrate the feasibility of implementing a harmonized process at multiple facilities for the manufacture of a complex cellular product. The quality systems and regulatory and operational strategies developed by the CIT Consortium yielded product lots that met the prespecified characteristics of safety, purity, potency, and identity and were successfully transplanted into 48 subjects. No adverse events attributable to the product and no cases of primary nonfunction were observed

Cell Replacement Therapy in Type 1 Diabetes

Cell Replacement Therapy in Type 1 Diabete

The Effect of Recovery Warm-up Time Following Cold Storage on the Dynamic Glucose-stimulated Insulin Secretion of Isolated Human Islets

Standardized islet characterization assays that can provide results in a timely manner are essential for successful islet cell transplantation. A critical component of islet cell quality is β-cell function, and perifusion-based assessments of dynamic glucose-stimulated insulin secretion (GSIS) are the most informative method to assess this, as they provide the most complex in vitro evaluation of GSIS. However, protocols used vary considerably among centers and investigators as they often use different low- and high-glucose concentrations, exposure-times, flow-rates, oxygen concentrations, islet numbers, analytical methods, measurement units, and instruments, which result in different readouts and make comparisons across platforms difficult. Additionally, the conditions of islet storage and shipment prior to assessment may also affect islet function. Establishing improved standardized protocols for perifusion GSIS assays should be an integral part of the ongoing effort to increase the rigor of human islet studies. Here, we performed detailed evaluation of GSIS of human islets using a fully automated multichannel perifusion instrument following various warm-up recovery times after cold storage that corresponds to current shipping conditions (8°C). We found that recovery times shorter than 18 h (overnight) resulted in impaired insulin secretion. While the effects were relatively moderate on second-phase insulin secretion, first-phase peaks were restored only following 18-h incubation. Hence, the biphasic profile of dynamic GSIS was considerably affected when islets were not allowed to recover for a sufficient time after being maintained in cold. Accordingly, while cold storage might improve islet cell survival during shipment and prolong the length of culture, functional assessments should be performed only after allowing for at least overnight recovery at physiological temperatures

Expression of c-fos Protein in Rat Brain Elicited by Electrical and Chemical Stimulation of the Hypothalamic Paraventricular Nucleus

The functional connectivity of the paraventricular nucleus of the hypothalamus (PVN) was studied by assessing the expression of the immediate early gene, c-fos, after unilateral stimulation of this structure in urethane-anesthetized rats. Electrical stimulation for 1 h (10 s on, 10 s off; 15-40 µA at 20 Hz) was accompanied by increases in mean arterial pressure (13-29 mm Hg). In these animals, ipsilateral increases in numbers of neurons with Fos-like immunoreactivity (FLI) were immunohistochemically demonstrated in the insular cortex, lateral septum, medial amygdala, hypothalamus, lateral division of the parabrachial nucleus (PBN) of the pons and the nucleus of the tractus solitarius (NTS) and ventrolateral medulla (VLM). Numbers of cells with FLI were quantitated in five areas known for their roles in autonomic function: arcuate nucleus, ventromedial hypothalamus, lateral PBN, NTS (at three levels) and VLM (caudal and rostral). In each case, stimulation of the PVN led to significant differences in number of neurons with FLI on the side ipsilateral to the stimulation compared to the contralateral side. To eliminate effects associated with stimulation of fibers of passage in the vicinity of the PVN, the results after electrical stimulation were compared to those obtained in animals in which the PVN was chemically stimulated unilaterally with the excitatory amino acid L-glutamate (5 one-minute infusions of 50 nl, 0.5 M glutamate over 1 h). Mean arterial pressure was increased after each injection (7–13 mm Hg), and significant differences in numbers of neurons with FLI between sides were maintained in all five areas except the NTS caudal to, and at, the level of the area postrema. An increase in neurons with FLI in the piriform cortex of all animals including controls may be due to injury-induced activation of target neurons from the PVN. These data illustrate that electrical and chemical stimulation of the PVN leads to simultaneous activation of neurons in many targets. All of the target areas studied receive direct projections from the PVN, although multisynaptic projections may also contribute to activation of target neurons

Fully Automated Islet Cell Counter (ICC) for the Assessment of Islet Mass, Purity, and Size Distribution by Digital Image Analysis

For isolated pancreatic islet cell preparations, it is important to be able to reliably assess their mass and quality, and for clinical applications, it is part of the regulatory requirement. Accurate assessment, however, is difficult because islets are spheroid-like cell aggregates of different sizes (<50 to 500 μm) resulting in possible thousandfold differences between the mass contribution of individual particles. The current standard manual counting method that uses size-based group classification is known to be error prone and operator dependent. Digital image analysis (DIA)-based methods can provide less subjective, more reproducible, and better-documented islet cell mass (IEQ) estimates; however, so far, none has become widely accepted or used. Here we present results obtained using a compact, self-contained islet cell counter (ICC3) that includes both the hardware and software needed for automated islet counting and requires minimal operator training and input; hence, it can be easily adapted at any center and could provide a convenient standardized cGMP-compliant IEQ assessment. Using cross-validated sample counting, we found that for most human islet cell preparations, ICC3 provides islet mass (IEQ) estimates that correlate well with those obtained by trained operators using the current manual SOP method ( r = 0.78, slope = 1.02). Variability and reproducibility are also improved compared to the manual method, and most of the remaining variability (CV = 8.9%) results from the rearrangement of the islet particles due to movement of the sample between counts. Characterization of the size distribution is also important, and the present digitally collected data allow more detailed analysis and coverage of a wider size range. We found again that for human islet cell preparations, a Weibull distribution function provides good description of the particle size

Targeted bone marrow radioablation with 153Samarium-lexidronam promotes allogeneic hematopoietic chimerism and donor-specific immunologic hyporesponsiveness

Transplantation tolerance, defined as acceptance of a graft by an otherwise fully immunocompetent host, has been an elusive goal. Although robust tolerance has been achieved by the induction of stable hematopoietic chimerism after bone marrow transplantation, lethal or sublethal radiation conditioning used to induce long-term chimerism precludes its clinical use. We studied whether targeted delivery of radiation to bone marrow could allow for bone marrow cell (BMC) engraftment, chimerism, and donor-specific tolerance in the absence of the side effects associated with external irradiation. We administered a radioactive bone-seeking compound (Samarium-Lexidronam, Quadramet, Berlex Laboratories, Wayne, NJ) together with transient T-cell costimulatory blockade to recipient mice. Allogeneic BMCs were given 7 or 14 days after preconditioning. Costimulatory blockade was obtained by the use of an anti-CD154 antibody for 4 weeks. Chimerism was assessed by flow cytometry. Mice then received donor-specific and third-party skin grafts. Graft survival was analyzed with mechanisms of donor-specific hyporesponsiveness. High levels of stable chimerism across an allogeneic barrier were achieved in mice by a single administration of Samarium-Lexidronam, transient T-cell costimulatory blockade, and BMC transplantation. A large percentage of chimeric animals retained donor-derived skin grafts for more than 120 days without requiring additional immunosuppression, suggesting that harsh cytotoxic preconditioning is not necessary to achieve stable chimerism and donor specific hyporesponsiveness. Analysis of the T-cell repertoire in chimeras indicates T-cell deletional mechanisms. These data broaden the potential use of BMC transplantation for tolerance induction and argue for its potential in treating autoimmune diseases