Despite the potent immunosuppressive activity that mesenchymal stem cells (MSCs) display in vitro, recent clinical trial results are disappointing, suggesting that MSC viability and/or function are greatly reduced after infusion. In this report, we demonstrated that human MSCs activated complement of the innate immunity after their contact with serum. Although all 3 known intrinsic cell-surface complement regulators were present on MSCs, activated complement overwhelmed the protection of these regulators and resulted in MSCs cytotoxicity and dysfunction. In addition, autologous MSCs suffered less cellular injury than allogeneic MSCs after contacting serum. All 3 complement activation pathways were involved in generating the membrane attack complex to directly injure MSCs. Supplementing an exogenous complement inhibitor, or up-regulating MSC expression levels of CD55, one of the cell-surface complement regulators, helped to reduce the serum-induced MSC cytotoxicity. Finally, adoptively transferred MSCs in complement deficient mice or complement-depleted mice showed reduced cellular injury in vivo compared with those in wild type mice. These results indicate that complement is integrally involved in recognizing and injuring MSCs after their infusion, suggesting that autologous MSCs may have advantages over allogeneic MSCs, and that inhibiting complement activation could be a novel strategy to improve existing MSC-based therapies. (Blood. 2012;120(17):3436-3443) IntroductionMesenchymal stem cells (MSCs) are adult stem cells that can differentiate into many types of cells, and they are strongly immunosuppressive. 1 MSCs have great potential in inflammatory disease treatment and in regenerative medicine. 2 There have been approximately 150 MSC-based clinical trials registered in the www.clinicaltrials.gov website alone for the treatment of different diseases, including graft-versus-host disease (GVHD), Crohn disease, diabetes, multiple sclerosis (MS), myocardial infarction, and allograft rejection. It is widely believed in the field that MSCs are able to escape from the host immune surveillance; therefore, MSCs expanded from 1 donor can be used to treat other patients without being rejected. 2,3 However, despite their apparent immunosuppressive activity in vitro, results from recent MSCs clinical trials were not very encouraging. 4 One possible explanation for these disappointing results is that immediately after systemic infusion, these MSCs are recognized and injured by the host, leading to severely reduced activity in vivo.Complement is an important part of the innate immunity, which serves as the first shield against anything foreign, and complement components are abundant in the serum. 5 Complement can be activated through 3 different pathways: the classic, the alternative, and the lectin pathway. Activated complement recruits and activates leukocytes, enhances phagocytosis, and forms membrane attack complex (MAC) to directly injure the target cells. It has been established that complement plays an important rol...

Human mesenchymal stem cell (hMSC) derived extracellular vesicles (EVs) have shown therapeutic potential in recent studies. However, the corresponding therapeutic components are largely unknown, and scale-up production of hMSC EVs is a major challenge for translational applications. In the current study, hMSCs were grown as 3D aggregates under wave motion to promote EV secretion. Results demonstrate that 3D hMSC aggregates promote activation of the endosomal sorting complexes required for transport (ESCRT)-dependent and -independent pathways. mRNA sequencing revealed global transcriptome alterations for 3D hMSC aggregates. Compared to 2D-hMSC-EVs, the quantity of 3D-hMSC-EVs was enhanced significantly (by 2-fold), with smaller sizes, higher miR-21 and miR-22 expression, and an altered protein cargo (e.g., upregulation of cytokines and anti-inflammatory factors) uncovered by proteomics analysis, possibly due to altered EV biogenesis. Functionally, 3D-hMSC-EVs rejuvenated senescent stem cells and exhibited enhanced immunomodulatory potentials. In summary, this study provides a promising strategy for scalable production of high-quality EVs from hMSCs with enhanced therapeutic potential.

Chronic pain affects one in five of the general population and is the third most important cause of disability-adjusted life-years globally. Unfortunately, treatment remains inadequate due to poor efficacy and tolerability. There has been a failure in translating promising preclinical drug targets into clinic use. This reflects challenges across the whole drug development pathway, from preclinical models to trial design. Nociceptors remain an attractive therapeutic target: their sensitization makes an important contribution to many chronic pain states, they are located outside the blood–brain barrier, and they are relatively specific. The past decade has seen significant advances in the techniques available to study human nociceptors, including: the use of corneal confocal microscopy and biopsy samples to observe nociceptor morphology, the culture of human nociceptors (either from surgical or post-mortem tissue or using human induced pluripotent stem cell derived nociceptors), the application of high throughput technologies such as transcriptomics, the in vitro and in vivo electrophysiological characterization through microneurography, and the correlation with pain percepts provided by quantitative sensory testing. Genome editing in human induced pluripotent stem cell-derived nociceptors enables the interrogation of the causal role of genes in the regulation of nociceptor function. Both human and rodent nociceptors are more heterogeneous at a molecular level than previously appreciated, and while we find that there are broad similarities between human and rodent nociceptors there are also important differences involving ion channel function, expression, and cellular excitability. These technological advances have emphasized the maladaptive plastic changes occurring in human nociceptors following injury that contribute to chronic pain. Studying human nociceptors has revealed new therapeutic targets for the suppression of chronic pain and enhanced repair. Cellular models of human nociceptors have enabled the screening of small molecule and gene therapy approaches on nociceptor function, and in some cases have enabled correlation with clinical outcomes. Undoubtedly, challenges remain. Many of these techniques are difficult to implement at scale, current induced pluripotent stem cell differentiation protocols do not generate the full diversity of nociceptor populations, and we still have a relatively poor understanding of inter-individual variation in nociceptors due to factors such as age, sex, or ethnicity. We hope our ability to directly investigate human nociceptors will not only aid our understanding of the fundamental neurobiology underlying acute and chronic pain but also help bridge the translational gap.

Stem cells are promising cell sources for many biomedical applications including cell therapy, regenerative medicine, and drug discovery. However, the commonly used static tissue culture vessels can only generate a low number of cells. To provide an adequate number of stem cells for clinical applications, a scalable process based on bioreactors is needed. Stem cells can be either cultured as free cells/aggregates in suspension or as adherent cells on the solid substrates. Based on the cell property, different bioreactor configurations are developed to better expand stem cells while maintaining their differentiation capacity. In this review, several major types of bioreactor systems and their applications in stem cell engineering are discussed. Continued advancements in bioprocess and bioreactor research and development are important to engineer stem cells for their use in biomedical applications.

The diversity and complexity of the cancer transcriptome may contain transcripts unique to the tumor environment. Here, we report a LIN28B variant, LIN28B-TST, which is specifically expressed in hepatocellular carcinoma (HCC) and many other cancer types. Expression of LIN28B-TST is associated with significantly poor prognosis in HCC patients. LIN28B-TST initiates from a de novo alternative transcription initiation site that harbors a strong promoter regulated by NFYA but not c-Myc. Demethylation of the LIN28B-TST promoter might be a prerequisite for its transcription and transcriptional regulation. LIN28B-TST encodes a protein isoform with additional N-terminal amino acids and is critical for cancer cell proliferation and tumorigenesis. Our findings reveal a mechanism of LIN28B activation in cancer and the potential utility of LIN28B-TST for clinical purposes.