Ectopic Tissue Formation Research Articles

Subchronic intranasal lipopolysaccharide exposure induces pulmonary autoimmunity and glomerulonephritis in NZBWF1 mice

Lupus, a systemic autoimmune disease shaped by gene-environment interplay, often progresses to endstage renal failure. While subchronic systemic exposure to bacterial lipopolysaccharide (LPS) triggers autoimmunity and glomerulonephritis in lupus-prone mice, it is unknown if inhaling LPS, which is common in certain occupations, can similarly trigger lupus. Here we determined how subchronic intranasal (IN) LPS instillation influences autoimmunity and glomerulonephritis development in lupusprone NZBWF1 female mice. Briefly, mice were IN-instilled with vehicle or E. coli LPS (0.8 μg/g) twice weekly for 5 wk, followed by necropsy. For systemic comparison, additional cohorts of mice were injected with LPS intraperitoneally (IP) using identical doses/timing. Lungs were assessed for inflammatory and autoimmune responses and then related to systemic autoimmunity and glomerulonephritis. IN/LPS exposure induced in the lung: i) leukocyte infiltration, ii)mRNA signatures for cytokines, chemokines, IFN-regulated, and cell death-related genes, iii) ectopic lymphoid tissue formation, and iv)diverse IgM and IgG autoantibodies (AAbs). Pulmonary effects coincided with enlarged spleens, elevated plasma IgG AAbs, and inflamed IgG-containing kidney glomeruli. In contrast, IP/LPS treatment induced systemic autoimmunity and glomerulonephritis without pulmonary manifestations. Taken together, these preclinical findings suggest the lung could serve as a critical nexus for triggering autoimmunity by respirable LPS in genetically predisposed individuals.

Single-cell profiling indicates a high similarity between immune cells in the cerebrospinal fluid and in meningeal ectopic lymphoid tissue in experimental autoimmune encephalomyelitis.

B cell depleting anti-CD20 monoclonal antibodies (aCD20 mAbs) are highly effective in treatment of multiple sclerosis (MS) but fail to halt the formation of meningeal ectopic lymphoid tissue (mELT) in the murine model experimental autoimmune encephalomyelitis (EAE). While mELT can be examined in EAE, it is not accessible in vivo in MS patients. Our key objectives were to compare the immune cells in cerebrospinal fluid (CSF), which is accessible in patients, with those in mELT, and to study the effects of aCD20 mAbs on CSF and mELT in EAE. Applying single cell RNA sequencing, we compared gene expression profiles in immune cells from (1) CSF with mELT and (2) aCD20 mAbs treated with control treated mice in a spontaneous 2D2xTh EAE model. The immune cell composition in CSF and mELT was very similar. Gene expression profiles and pathway enrichment analysis revealed no striking differences between the two compartments. aCD20 mAbs led not only to a virtually complete depletion of B cells in the CSF but also to a reduction of naïve CD4+ T cells and marked increase of macrophages. No remarkable differences in regulated genes or pathways were observed. Our results suggest that immune cells in the CSF may serve as a surrogate for mELT in EAE. Future studies are required to confirm this in MS patients. The observed increase of macrophages in B cell depleted CSF is a novel finding and requires verification in CSF of aCD20 mAbs treated MS patients. Due to unresolved technical challenges, we were unable to study the effects of aCD20 mAbs on mELT. This should be addressed in future studies.

In vitro and in vivo evaluation of periosteum-derived cells and iPSC-derived chondrocytes encapsulated in GelMA for osteochondral tissue engineering.

Osteochondral defects are deep joint surface lesions that affect the articular cartilage and the underlying subchondral bone. In the current study, a tissue engineering approach encompassing individual cells encapsulated in a biocompatible hydrogel is explored in vitro and in vivo. Cell-laden hydrogels containing either human periosteum-derived progenitor cells (PDCs) or human induced pluripotent stem cell (iPSC)-derived chondrocytes encapsulated in gelatin methacryloyl (GelMA) were evaluated for their potential to regenerate the subchondral mineralized bone and the articular cartilage on the joint surface, respectively. PDCs are easily isolated and expanded progenitor cells that are capable of generating mineralized cartilage and bone tissue in vivo via endochondral ossification. iPSC-derived chondrocytes are an unlimited source of stable and highly metabolically active chondrocytes. Cell-laden hydrogel constructs were cultured for up to 28 days in a serum-free chemically defined chondrogenic medium. On day 1 and day 21 of the differentiation period, the cell-laden constructs were implanted subcutaneously in nude mice to evaluate ectopic tissue formation 4 weeks post-implantation. Taken together, the data suggest that iPSC-derived chondrocytes encapsulated in GelMA can generate hyaline cartilage-like tissue constructs with different levels of maturity, while using periosteum-derived cells in the same construct type generates mineralized tissue and cortical bone in vivo. Therefore, the aforementioned cell-laden hydrogels can be an important part of a multi-component strategy for the manufacturing of an osteochondral implant.

Crystalline silica-induced pulmonary inflammation and autoimmunity in mature adult NZBW/f1 mice: age-related sensitivity and impact of omega-3 fatty acid intervention

Objective Occupational exposure to respirable crystalline silica (cSiO2) has been linked to lupus development. Previous studies in young lupus-prone mice revealed that intranasal cSiO2 exposure triggered autoimmunity, preventable with docosahexaenoic acid (DHA). This study explores cSiO2 and DHA effects in mature lupus-prone adult mice, more representative of cSiO2-exposed worker age. Methods Female NZBWF1 mice (14-week old) were fed control (CON) or DHA-supplemented diets. After two weeks, mice were intranasally instilled saline (VEH) or 1 mg cSiO2 weekly for four weeks. Cohorts were then analyzed 1- and 5-weeks postinstillation for lung inflammation, cell counts, chemokines, histopathology, B- and T-cell infiltration, autoantibodies, and gene signatures, with results correlated to autoimmune glomerulonephritis onset. Results VEH/CON mice showed no pathology. cSiO2/CON mice displayed significant ectopic lymphoid tissue formation in lungs at 1 week, increasing by 5 weeks. cSiO2/CON lungs exhibited elevated cellularity, chemokines, CD3+ T-cells, CD45R + B-cells, IgG + plasma cells, gene expression, IgG autoantibodies, and glomerular hypertrophy. DHA supplementation mitigated all these effects. Discussion The mature adult NZBWF1 mouse used here represents a life-stage coincident with immunological tolerance breach and one that more appropriately represents the age (20–30 yr) of cSiO2-exposed workers. cSiO2-induced robust pulmonary inflammation, autoantibody responses, and glomerulonephritis in mature adult mice, surpassing effects observed previously in young adults. DHA at a human-equivalent dosage effectively countered cSiO2-induced inflammation/autoimmunity in mature mice, mirroring protective effects in young mice. Conclusion These results highlight life-stage significance in this preclinical lupus model and underscore omega-3 fatty acids’ therapeutic potential against toxicant-triggered autoimmune responses.

Mettl3 regulates hypertrophic differentiation of chondrocytes through modulating Dmp1 mRNA via Ythdf1-mediated m6A modification.

As the main cells in endochondral osteogenesis, chondrocytes have limited self-repair ability due to weak proliferation activity, low density, and dedifferentiation tendency. Here, a thorough inquiry about the effect and underlying mechanisms of methyltransferase like-3 (Mettl3) on chondrocytes was made. Functionally, it was indicated that Mettl3 promoted the proliferation and hypertrophic differentiation of chondrocytes. Mechanically, Dmp1 (dentin matrix protein 1) was proved to be the downstream direct target of Mettl3 for m6A modification to regulate the differentiation of chondrocytes through bioinformatics analysis and correlated experiments. The Reader protein Ythdf1 mediated Dmp1 mRNA catalyzed by Mettl3. In vivo, the formation of subcutaneous ectopic cartilage-like tissue further supported the in vitro results. In conclusion, the gene regulation of Mettl3/m6A/Ythdf1/Dmp1 axis in hypertrophic differentiation of chondrocytes for the development of endochondral osteogenesis may supply a promising treatment strategy for the repair and regeneration of bone defects.

AFF4 regulates cellular adipogenic differentiation via targeting autophagy

Transcriptional elongation is a universal and critical step during gene expression. The super elongation complex (SEC) regulates the rapid transcriptional induction by mobilizing paused RNA polymerase II (Pol II). Dysregulation of SEC is closely associated with human diseases. However, the physiological role of SEC during development and homeostasis remains largely unexplored. Here we studied the function of SEC in adipogenesis by manipulating an essential scaffold protein AF4/FMR2 family member 4 (AFF4), which assembles and stabilizes SEC. Knockdown of AFF4 in human mesenchymal stem cells (hMSCs) and mouse 3T3-L1 preadipocytes inhibits cellular adipogenic differentiation. Overexpression of AFF4 enhances adipogenesis and ectopic adipose tissue formation. We further generate Fabp4-cre driven adipose-specific Aff4 knockout mice and find that AFF4 deficiency impedes adipocyte development and white fat depot formation. Mechanistically, we discover AFF4 regulates autophagy during adipogenesis. AFF4 directly binds to autophagy-related protein ATG5 and ATG16L1, and promotes their transcription. Depleting ATG5 or ATG16L1 abrogates adipogenesis in AFF4-overepressing cells, while overexpression of ATG5 and ATG16L1 rescues the impaired adipogenesis in Aff4-knockout cells. Collectively, our results unveil the functional importance of AFF4 in regulating autophagy and adipogenic differentiation, which broaden our understanding of the transcriptional regulation of adipogenesis.

Fabrication of bone-derived decellularized extracellular matrix/ceramic-based biocomposites and their osteo/odontogenic differentiation ability for dentin regeneration.

The goal of this study was to fabricate bioactive cell‐laden biocomposites supplemented with bone‐derived decellularized extracellular matrix (dECM) with calcium phosphate ceramic, and to assess the effect of the biocomponents on the osteogenic and odontogenic differentiation of human dental pulp stem cells (hDPSCs). By evaluating the rheological properties and selecting printing parameters, mechanically stable cell‐laden 3D biocomposites with high initial cell‐viability (>90%) and reasonable printability (≈0.9) were manufactured. The cytotoxicity of the biocomposites was evaluated via MTT assay and nuclei/F‐actin fluorescent analyses, while the osteo/odontogenic differentiation of the hDPSCs was assessed using histological and immunofluorescent analyses and various gene expressions. Alkaline phosphate activity and alizarin red staining results indicate that the dECM‐based biocomposites exhibit significantly higher osteogenic activities, including calcification, compared to the collagen‐based biocomposites. Furthermore, immunofluorescence images and gene expressions demonstrated upregulation of dentin matrix acidic phosphoprotein‐1 and dentin sialophosphoprotein in the dECM‐based biocomposites, indicating acceleration of the odontogenic differentiation of hDPSCs in the printed biocomposites. The hDPSC‐laden biocomposite was implanted into the subcutaneous region of mice, and the biocomposite afforded clear induction of osteo/odontogenic ectopic hard tissue formation 8 weeks post‐transplantation. From these results, we suggest that the hDPSC‐laden biocomposite is a promising biomaterial for dental tissue engineering.

INSPECT: A Retrospective Study to Evaluate Long-term Effectiveness and Safety of Darvadstrocel in Patients With Perianal Fistulizing Crohn's Disease Treated in the ADMIRE-CD Trial.

The efficacy of a single administration of darvadstrocel (expanded allogeneic adipose-derived mesenchymal stem cells) for treating complex perianal fistulas in patients with Crohn's disease was demonstrated in a randomized, double-blind trial (ADMIRE-CD [Adipose Derived Mesenchymal Stem Cells for Induction of Remission in Perianal Fistulizing Crohn\'s Disease] trial). The current chart review study (INSPECT [A retrospectIve chart review study evaluatINg the longer-term effectiveneSs of darvadstrocel in PatiEnts who CompleTed ADMIRE-CD]) evaluated the longer-term effectiveness and safety of darvadstrocel. Eligible patients had completed at least 52 weeks in the ADMIRE-CD trial. Data on clinical remission and fistula relapse outcomes were collected retrospectively at 104 and 156 weeks after treatment. Adverse events of special interest (tumorigenicity and ectopic tissue formation) were collected up to 208 weeks after treatment. Eighty-nine patients were included (43 darvadstrocel patients, 46 control subjects). At 52, 104, and 156 weeks posttreatment, clinical remission was observed in 29 (67.4%) of 43, 23 (53.5%) of 43, and 23 (53.5%) of 43 darvadstrocel-treated patients, compared with 24 (52.2%) of 46, 20 (43.5%) of 46, and 21 (45.7%) of 46 control subjects, respectively. In patients with clinical remission at week 52, this remission was sustained at 104 and 156 weeks after treatment in 19 (65.5%) of 29 and 16 (55.2%) of 29 darvadstrocel-treated patients and in 17 (70.8%) of 24 and 13 (54.2%) of 24 control subjects, respectively. Time to fistula relapse and incidence of fistula relapse or new fistula occurrence were not significantly different between groups. Tumorigenicity was reported for 1 (2.2%) patient in the control group (malignant epidermoid carcinoma). No ectopic tissue formation was reported. Real-world follow-up of patients from the ADMIRE-CD trial indicates that clinical remission of complex perianal fistulas can be sustained in the long term irrespective of whether it is achieved through darvadstrocel administration or maintenance treatment regimens and confirms a favorable long-term safety profile of darvadstrocel.

P603 INSPIRE: 6-month interim analysis from an observational post-marketing registry on the effectiveness and safety of darvadstrocel in patients with Crohn’s disease and complex perianal fistulas

Abstract Background Darvadstrocel (DVS) is a suspension of expanded allogeneic adipose-derived mesenchymal (stromal) stem cells approved for the treatment of complex Crohn’s perianal fistulas (complex CPF). The INSPIRE (EUPAS24267) registry is a European observational, post-approval study designed to evaluate the real-world effectiveness and safety of DVS in patients with complex CPF for up to 36 months after DVS administration. Here we report clinical and safety outcomes from the first interim analysis at 6 months post treatment. Methods Patients with complex CPF treated with DVS according to local standard practice were eligible to enrol. Primary outcomes were clinical response (defined as closure despite gentle finger compression of ≥50% of external openings treated with DVS that were draining at baseline) and clinical remission (defined as closure despite gentle finger compression of all external openings treated with DVS that were draining at baseline). Outcomes were evaluated in the All Treated (AT) and Treated Per Protocol (PP) cohorts (Table 1) 6 months post treatment in patients who had received a single administration of DVS. The number and nature of adverse events (AEs), serious AEs (SAEs) are reported for the AT cohort. Results As of September 2021, 230 patients had enrolled, 225 met the inclusion criteria and 205 had complete treatment data and comprised the AT cohort, of these, 176 were treated per protocol. A total of 138 (AT) and 120 (PP) patients were 6 months post treatment, 66% (92/138) and 58% (69/120) had a 6-month visit, of which, 85% (78/92) and 100% (69/69) had evaluable data at Month 6, respectively. Baseline demographics and clinical and fistula characteristics are shown in Tables 1 and 2. In the AT and PP cohorts, respectively, 73% (57/78) and 74% (51/69) of patients had a clinical response, and 65% (51/78) and 65% (45/69) had clinical remission. Changes in Crohn’s disease (CD) activity (assessed with the Harvey–Bradshaw Index) post treatment were minimal, and most patients had no change in CD activity from baseline. In the AT cohort, 20% (41/205) of patients had ≥1 AE and 9.3% (19/205) had ≥1 SAE. There were no reports of ectopic tissue formation and no deaths. Conclusion This is the first real-world, multi-centre study reporting the effectiveness and safety data after DVS treatment for complex CPF. These interim analysis data are consistent with the pivotal ADMIRE-CD study in terms of effectiveness and safety. The ongoing status of the registry and small number of evaluable patients mean these data should be interpreted with caution, pending future analyses in larger numbers of patients with longer follow-up times.

Stem cells or their exosomes: which is preferred in COVID-19 treatment?

It only took 8 months for the pneumonia caused by a previously unknown coronavirus to turn into a global pandemic of unprecedentedly far-reaching implications. Failure of the already discovered treatment measures opened up a new opportunity to evaluate the potentials of mesenchymal stem cells and their extracellular vesicles (EVs), exosomes in particular. Eventually, the initial success experienced after the use of MSCs in treating the new pneumonia by Lnge and his team backed up the idea of MSC-based therapies and pushed them closer to becoming a reality. However, MSC-related concerns regarding safety such as abnormal differentiation, spontaneous malignant and the formation of ectopic tissues have triggered the replacement of MSCs by their secreted exosomes. The issue has been further strengthened by the fact that the exosomes leave similar treatment impacts when compared to their parental cells. In recent years, much attention has been paid to the use of MSC-derived exosomes in the treatment of a variety of diseases. With a primary focus on COVID-19 and its current treatment methods, the present review looks into the potentials of MSCs and MSC-derived exosomes in battling the ongoing pandemic. Finally, the research will draw an analogy between exosomes and their parental cells, when it comes to the progresses and challenges in using exosomes as a large-scale treatment method.

Siponimod Inhibits the Formation of Meningeal Ectopic Lymphoid Tissue in Experimental Autoimmune Encephalomyelitis

Background and ObjectivesTo investigate whether the formation or retention of meningeal ectopic lymphoid tissue (mELT) can be inhibited by the sphingosine 1-phosphate receptor 1,5 modulator siponimod (BAF312) in a murine model of multiple sclerosis (MS).MethodsA murine spontaneous chronic experimental autoimmune encephalomyelitis (EAE) model, featuring meningeal inflammatory infiltrates resembling those in MS, was used. To prevent or treat EAE, siponimod was administered daily starting either before EAE onset or at peak of disease. The extent and cellular composition of mELT, the spinal cord parenchyma, and the spleen was assessed by histology and immunohistochemistry.ResultsSiponimod, when applied before disease onset, ameliorated EAE. This effect was also present, although less prominent, when treatment started at peak of disease. Treatment with siponimod resulted in a strong reduction of the extent of mELT in both treatment paradigms. Both B and T cells were diminished in the meningeal compartment.DiscussionBeneficial effects on the disease course correlated with a reduction in mELT, suggesting that inhibition of mELT may be an additional mechanism of action of siponimod in the treatment of EAE. Further studies are needed to establish causality and confirm this observation in MS.

MiRNA induced co-differentiation and cross-talk of adipose tissue-derived progenitor cells for 3D heterotypic pre-vascularized bone formation

Engineered bone grafts require a vascular network to supply cells with oxygen, nutrients and remove waste. Using heterotypic mature cells to create these grafts in vivo has resulted in limited cell density, ectopic tissue formation and disorganized tissue. Despite evidence that progenitor cell aggregates, such as progenitor spheroids, are a potential candidate for fabrication of native-like pre-vascularized bone tissue, the factors dictating progenitor co-differentiation to create heterotypic pre-vascularized bone tissue remains poorly understood. In this study, we examined a three-dimensional heterotypic pre-vascularized bone tissue model, using osteogenic and endotheliogenic progenitor spheroids induced by miR-148b and miR-210 mimic transfection, respectively. Spheroids made of transfected cells were assembled into heterotypic structures to determine the impact on co-differentiation as a function of micro-RNA (miRNA) mimic treatment group and induction time. Our results demonstrated that miRNAs supported the differentiation in heterotypic structures, and that developing heterotypic structures is determined in part by progenitor maturity, as confirmed by gene and protein markers of osteogenic and endotheliogenic differentiation and the mineralization assay. As a proof of concept, miRNA-transfected spheroids were also bioprinted using aspiration-assisted bioprinting and organized into hollow structures to mimic the Haversian canal. Overall, the presented approach could be useful in fabrication of vascularized bone tissue using spheroids as building blocks.

Bcl6 controls meningeal Th17–B cell interaction in murine neuroinflammation

Ectopic lymphoid tissue containing B cells forms in the meninges at late stages of human multiple sclerosis (MS) and when neuroinflammation is induced by interleukin (IL)-17 producing T helper (Th17) cells in rodents. B cell differentiation and the subsequent release of class-switched immunoglobulins have been speculated to occur in the meninges, but the exact cellular composition and underlying mechanisms of meningeal-dominated inflammation remain unknown. Here, we performed in-depth characterization of meningeal versus parenchymal Th17-induced rodent neuroinflammation. The most pronounced cellular and transcriptional differences between these compartments was the localization of B cells exhibiting a follicular phenotype exclusively to the meninges. Correspondingly, meningeal but not parenchymal Th17 cells acquired a B cell-supporting phenotype and resided in close contact with B cells. This preferential B cell tropism for the meninges and the formation of meningeal ectopic lymphoid tissue was partially dependent on the expression of the transcription factor Bcl6 in Th17 cells that is required in other T cell lineages to induce isotype class switching in B cells. A function of Bcl6 in Th17 cells was only detected in vivo and was reflected by the induction of B cell-supporting cytokines, the appearance of follicular B cells in the meninges, and of immunoglobulin class switching in the cerebrospinal fluid. We thus identify the induction of a B cell-supporting meningeal microenvironment by Bcl6 in Th17 cells as a mechanism controlling compartment specificity in neuroinflammation.

Immune checkpoints and the multiple faces of B cells in systemic lupus erythematosus.

B-lymphocytes are crucial in the pathogenesis of systemic lupus erythematosus (SLE), including autoantibody production, antigen presentation, co-stimulation, and cytokine secretion. Co-stimulatory and co-inhibitory molecules control interactions between B and T cells during an inflammatory response, which is essential for an appropriate host protection and maintenance of self-tolerance. Here, we review recent findings about checkpoint molecules and SLE B cells including their potential therapeutic implications and experiences from clinical trials. Most prominent checkpoint molecules involved in pathologic B and T cell interaction in SLE are CD40/CD40L and inducible co-stimulator/ICOSL, both also intimately involved in the formation of germinal centers and ectopic lymphoid tissue. Dysregulations of inhibitory checkpoint molecules, like programmed death-1/programmed death-ligand 1 and B- and T-lymphocyte attenuator have been suggested to impair B cell functions in SLE recently. Accumulating evidence indicates that dampening immune responses by either blocking co-activating signals or enhancing co-inhibitory signals in different cell types is a promising approach to treat autoimmune diseases to better control active disease but may also allow resolution of chronic autoimmunity.

Repeated intra-articular administration of equine allogeneic peripheral blood-derived mesenchymal stem cells does not induce a cellular and humoral immune response in horses

ObjectiveThe use of mesenchymal stem cells (MSCs) for the treatment of equine joint disease is widely investigated because of their regenerative and immunomodulatory potential. Allogeneic MSCs provide a promising alternative to autologous MSCs, since the former are immediately available and enable a thorough donor screening. However, questions have been raised concerning the immunogenic potential of allogeneic MSCs, especially after repeated administration. MethodsCurrent retrospective study assessed the cellular and humoral immunogenicity of ten jumping and dressage horses with naturally occurring degenerative joint disease which were treated 3 times intra-articularly with a 1 mL stem cell suspension containing 1.4–2.5 million chondrogenic induced equine allogeneic peripheral blood-derived MSCs (ciMSCs) combined with 1 mL equine allogeneic plasma. Stem cells from 2 donor horses were used. Horses were clinically evaluated for joint effusion, presence of pain to palpation and skin surface temperature at the local injection site, joint range of motion, occurrence of adverse events and the presence of ectopic tissue. The cellular immune response was analyzed using a modified mixed lymphocyte reaction and the humoral immune response was investigated using a flow cytometric crossmatch assay by which the presence of alloantibodies against the ciMSCs was evaluated. Presence of anti-bovine serum albumin antibodies was detected via ELISA. ResultsClinical evaluation of the horses revealed no serious adverse effects or suspected adverse drug reactions and no ectopic tissue formation at the local injection site or in other areas of the body. Generally, repeated administration led to a decrease of horses with joint effusion of the affected joint. Pain to palpation, skin surface temperature and joint range of motion did not increase or even decreased after treatment administration.Allogeneic ciMSCs did not induce a cellular immune response and no alloantibodies were detected in the recipients’ serum, regardless the presence of BSA antibodies in 70 % of the horses. ConclusionRepeated intra-articular injections with allogeneic equine ciMSCs did not elicit clinically relevant adverse events. Furthermore, current study indicates the absence of a cellular or a humoral immune response following repeated intra-articular injections.

Who am I? (re-)Defining fibroblast identity and immunological function in the age of bioinformatics.

Who am I? (re-)Defining fibroblast identity and immunological function in the age of bioinformatics.

Calcium Chelidonate: Semi-Synthesis, Crystallography, and Osteoinductive Activity In Vitro and In Vivo.

Calcium chelidonate [Ca(ChA)(H2O)3]n was obtained by semi-synthesis using natural chelidonic acid. The structure of the molecular complex was determined by X-ray diffraction analysis. The asymmetric unit of [Ca(ChA)(H2O)3]n includes chelidonic acid coordinated through three oxygen atoms, and three water ligands. The oxygen atoms of acid and oxygen atoms of water from each asymmetric unit are also coordinated to the calcium of another one, forming an infinite linear complex. Calcium geometry is close to the trigonal dodecahedron (D2d). The intra-complex hydrogen bonds additionally stabilize the linear species, which are parallel to the axis. In turn the linear species are packed into the 3D structure through mutual intercomplex hydrogen bonds. The osteogenic activity of the semi-synthetic CaChA was studied in vitro on 21-day hAMMSC culture and in vivo in mice using ectopic (subcutaneous) implantation of CaP-coated Ti plates saturated in vitro with syngeneic bone marrow. The enhanced extracellular matrix ECM mineralization in vitro and ectopic bone tissue formation in situ occurred while a water solution of calcium chelidonate at a dose of 10 mg/kg was used. The test substance promotes human adipose-derived multipotent mesenchymal stromal/stem cells (hAMMSCs), as well as mouse MSCs to differentiate into osteoblasts in vitro and in vivo, respectively. Calcium chelidonate is non-toxic and can stimulate osteoinductive processes.

The Multiple Roles of B Cells in the Pathogenesis of Sjögren's Syndrome.

Primary Sjögren’s syndrome (pSS) is a chronic autoimmune disease characterized by lymphocytic infiltration and tissue destruction of exocrine glands such as salivary glands. Although the formation of ectopic lymphoid tissue in exocrine glands and overproduction of autoantibodies by autoreactive B cells highlight the critical involvement of B cells in disease development, the precise roles of various B cell subsets in pSS pathogenesis remain partially understood. Current studies have identified several novel B cell subsets with multiple functions in pSS, among which autoreactive age-associated B cells, and plasma cells with augmented autoantibody production contribute to the disease progression. In addition, tissue-resident Fc Receptor-Like 4 (FcRL4)+ B cell subset with enhanced pro-inflammatory cytokine production serves as a key driver in pSS patients with mucosa-associated lymphoid tissue (MALT)-lymphomas. Recently, regulatory B (Breg) cells with impaired immunosuppressive functions are found negatively correlated with T follicular helper (Tfh) cells in pSS patients. Further studies have revealed a pivotal role of Breg cells in constraining Tfh response in autoimmune pathogenesis. This review provides an overview of recent advances in the identification of pathogenic B cell subsets and Breg cells, as well as new development of B-cell targeted therapies in pSS patients.

Anti-CD20 Depletes Meningeal B Cells but Does Not Halt the Formation of Meningeal Ectopic Lymphoid Tissue.

ObjectiveTo investigate whether anti-CD20 B-cell-depleting monoclonal antibodies (ɑCD20 mAbs) inhibit the formation or retention of meningeal ectopic lymphoid tissue (mELT) in a murine model of multiple sclerosis (MS).MethodsWe used a spontaneous chronic experimental autoimmune encephalomyelitis (EAE) model of mice with mutant T-cell and B-cell receptors specific for myelin oligodendrocyte glycoprotein (MOG), which develop meningeal inflammatory infiltrates resembling those described in MS. ɑCD20 mAbs were administered in either a preventive or a treatment regimen. The extent and cellular composition of mELT was assessed by histology and immunohistochemistry.ResultsɑCD20 mAb, applied in a paradigm to either prevent or treat EAE, did not alter the disease course in either condition. However, ɑCD20 mAb depleted virtually all B cells from the meningeal compartment but failed to prevent the formation of mELT altogether. Because of the absence of B cells, mELT was less densely populated with immune cells and the cellular composition was changed, with increased neutrophil granulocytes.ConclusionsThese results demonstrate that, in CNS autoimmune disease, meningeal inflammatory infiltrates may form and persist in the absence of B cells. Together with the finding that ɑCD20 mAb does not ameliorate spontaneous chronic EAE with mELT, our data suggest that mELT may have yet unknown capacities that are independent of B cells and contribute to CNS autoimmunity.

Immunological aspects of RPE cell transplantation.

Retinal pigment epithelial (RPE) cells have several functions, including support of the neural retina and choroid in the eye and immunosuppression. Cultured human RPE cells directly suppress inflammatory immune cells. For instance, they directly suppress the activation of T cells in vitro. In contrast, transplanted allogeneic human RPE cells are rejected by bystander immune cells such as T cells in vivo. Recently, human embryonic stem cell-derived RPE cells have been used in several clinical trials, and human induced pluripotent stem cell (iPSC)-RPE cells have also been tested in our clinical study in patients with retinal degeneration. Major safety concerns after stem cell-based transplantation surgery include hyper-proliferation, tumorigenicity, or ectopic tissue formation, but these events have currently not been seen in any of these patients. However, if RPE cells are allogeneic, there are concerns about immune rejection issues that have been raised in previous clinical trials. We therefore performed a preclinical study of allogeneic iPSC-RPE cell transplantation in animal rejection models. We then conducted autogenic or allogeneic iPSC-RPE cell transplantation in clinical studies of patients with age-related macular degeneration. In this review, we focus on immunological studies of RPE cells, including iPSC-derived cells. iPSC-RPE cells have unique inflammatory (immunosuppressive and immunogenic) characteristics like primary cultured RPE cells. The purpose of this review is to summarize the current findings obtained from preclinical (basic research) and clinical studies in iPSC-RPE cell transplantation, especially the immunological aspects.