Human iPSC-derived midbrain organoids functionally integrate into striatum circuits and restore motor function in a mouse model of Parkinson's disease.

Rationale: Parkinson's disease (PD) is a prevalent neurodegenerative disorder that is characterized by degeneration of dopaminergic neurons (DA) at the substantia nigra pas compacta (SNpc). Cell therapy has been proposed as a potential treatment option for PD, with the aim of replenishing the lost DA neurons and restoring motor function. Fetal ventral mesencephalon tissues (fVM) and stem cell-derived DA precursors cultured in 2-dimentional (2-D) culture conditions have shown promising therapeutic outcomes in animal models and clinical trials. Recently, human induced pluripotent stem cells (hiPSC)-derived human midbrain organoids (hMOs) cultured in 3-dimentional (3-D) culture conditions have emerged as a novel source of graft that combines the strengths of fVM tissues and 2-D DA cells. Methods: 3-D hMOs were induced from three distinct hiPSC lines. hMOs at various stages of differentiation were transplanted as tissue pieces into the striatum of naïve immunodeficient mouse brains, with the aim of identifying the most suitable stage of hMOs for cellular therapy. The hMOs at Day 15 were determined to be the most appropriate stage and were transplanted into a PD mouse model to assess cell survival, differentiation, and axonal innervation in vivo. Behavioral tests were conducted to evaluate functional restoration following hMO treatment and to compare the therapeutic effects between 2-D and 3-D cultures. Rabies virus were introduced to identify the host presynaptic input onto the transplanted cells. Results: hMOs showed a relatively homogeneous cell composition, mostly consisting of dopaminergic cells of midbrain lineage. Analysis conducted 12 weeks post-transplantation of day 15 hMOs revealed that 14.11% of the engrafted cells expressed TH+ and over 90% of these cells were co-labeled with GIRK2+, indicating the survival and maturation of A9 mDA neurons in the striatum of PD mice. Transplantation of hMOs led to a reversal of motor function and establishment of bidirectional connections with natural brain target regions, without any incidence of tumor formation or graft overgrowth. Conclusion: The findings of this study highlight the potential of hMOs as safe and efficacious donor graft sources for cell therapy to treat PD.

MicroRNAs of extracellular vesicles derived from mesenchymal stromal cells alleviate inflammation in dry eye disease by targeting the IRAK1/TAB2/NF-κB pathway.

PURPOSE: To investigate the efficacy and mechanisms of human umbilical cord-derived MSC-derived extracellular vesicles (hucMSC-EVs) in a mouse model of desiccation-induced dry eye disease (DED). METHODS: hucMSC-EVs were enriched by ultracentrifugation. The DED model was induced by desiccating environment combined with scopolamine administration. The DED mice were divided into the hucMSC-EVs group, fluorometholone (FML) group, PBS group, and blank control group. Tear secretion, corneal fluorescein staining, the cytokine profiles in tears and goblet cells, TUNEL-positive cell, and CD4+ cells were examined to assess therapeutic efficiency. The miRNAs in the hucMSC-EVs were sequenced, and the top 10 were used for miRNA enrichment analysis and annotation. The targeted DED-related signaling pathway was further verified by using RT‒qPCR and western blotting. RESULTS: Treatment with hucMSC-EVs increased the tear volume and maintained corneal integrity in DED mice. The cytokine profile in the tears of the hucMSC-EVs group presented with a lower level of proinflammatory cytokines than PBS group. Moreover, hucMSC-EVs treatment increased goblet cell density and inhibited cell apoptosis and CD4+ cell infiltration. Functional analysis of the top 10 miRNAs in hucMSC-EVs showed a high correlation with immunity. Among them, miR-125 b, let-7b, and miR-6873 were conserved between humans and mice and were associated with the IRAK1/TAB2/NF-κB pathway that was activated in DED. Furthermore, IRAK1/TAB2/NF-κB pathway activation and the abnormal expression of IL-4, IL-8, IL-10, IL-13, IL-17, and TNF-α were reversed by hucMSC-EVs. CONCLUSIONS: hucMSCs-EVs alleviate DED signs, suppress inflammation and restore homeostasis of the corneal surface by multitargeting the IRAK1/TAB2/NF-κB pathway via certain miRNAs.

Probiotic Formulation VSL#3 Interacts with Mesenchymal Stromal Cells To Protect Dopaminergic Neurons via Centrally and Peripherally Suppressing NOD-Like Receptor Protein 3 Inflammasome-Mediated Inflammation in Parkinson's Disease Mice.

Systemic immunomodulation is increasingly recognized among the beneficial effects of mesenchymal stromal cells (MSCs) in treatment of Parkinson's disease (PD), while the underlying mechanism is not fully understood. With the growing popularity of using probiotics as an adjuvant approach in PD treatment, concerns about the added effects of probiotics have been raised. In addition to the molecular mechanism mediating the neuroprotective effects of MSCs, the combined effects of a probiotic formulation, VSL#3, and MSC infusion were also evaluated in PD mice. The animals were weekly treated with human MSCs (hMSCs) via the tail vein, VSL#3 via the gastrointestinal tract, or their combination six times. hMSCs, VSL#3 alone, and their combination markedly ameliorated the decreased striatal dopamine content, loss of dopaminergic neurons in the substantia nigra, increased levels of proinflammatory cytokines in serum, as well as tumor necrosis factor alpha (TNF-α) and interleukin-1ß (IL-1ß) mRNAs in striatum and peripheral tissues induced by MPTP. Furthermore, hMSCs, VSL#3, and their combination notably downregulated mRNA expression of NOD-like receptor protein 3 (NLRP3) and caspase-1 in brain and peripheral tissues of PD mice. These results suggest that hMSCs, VSL#3, and their combination prevent neurodegenerative changes in PD mice via anti-inflammatory activities in both the central and peripheral systems, possibly through suppressing the NLRP3 inflammasome. Moreover, two-way analysis of variance (ANOVA) indicated that VSL#3 interacts with hMSCs to attenuate neurodegeneration and inhibit NLRP3 inflammasome-mediated inflammation without altering the effects of hMSCs. Major findings of our study support the usage of probiotic formulation VSL#3 as an adjuvant therapy to hMSC infusion in PD treatment. IMPORTANCE This study provides evidence for the neuroprotective activities of human umbilical cord MSCs from the aspect of anti-inflammation actions. hMSCs inhibit the NLRP3 inflammasome and MPTP-induced inflammation in both brain and periphery to relieve the degenerative changes in dopaminergic neurons in PD mice. Furthermore, as an additional therapeutic agent, probiotic formulation VSL#3 interacts with hMSCs in suppressing the NLRP3 inflammasome as well as the central and peripheral anti-inflammatory effects to exert neuroprotective actions in PD mice without altering the actions of hMSCs, suggesting the potential of VSL#3 as an adjuvant therapy in PD treatment. The findings of the present study give a further understanding of the anti-inflammatory activity and the molecular mechanism for the beneficial effects of MSCs as well as the potential application of probiotic formulation as an adjuvant approach to MSC therapy in PD treatment.

Neuroprotective effects of human umbilical cord mesenchymal stromal cells in PD mice via centrally and peripherally suppressing NLRP3 inflammasome-mediated inflammatory responses.

Mesenchymal stromal cells (MSCs) exhibit beneficial anti-inflammatory effects against Parkinson's disease (PD) via immunomodulatory actions. However, the underlying molecular mechanism remains unclear. This study aimed to investigate the potential neuroprotective effects of MSCs and the possible mechanisms involved by infusing human umbilical cord MSCs (hMSCs) in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mature male C57BL/6 mice. Subsequently, the striatal content of dopamine (DA) and its metabolites, tyrosine hydroxylase (TH)-positive neurons and activated microglia, circulating inflammatory cytokines, gene expression of cytokines, and NOD-like receptor protein 3 (NLRP3) inflammasome molecules were measured using high-performance liquid chromatography, flow cytometry, immunohistochemistry, immunofluorescent staining, and real-time polymerase chain reaction assays, respectively. Infused hMSCs markedly ameliorated the reduction in striatal DA and loss of TH-positive neurons in the substantia nigra of PD mice. The MPTP-induced activation of microglia and increase in tumor necrosis factor-α and interleukin-1ß mRNA expression in the striatum were also attenuated by hMSCs. Furthermore, hMSCs completely reversed the elevated pro-inflammatory cytokine levels in the serum and mRNA expression of cytokines in the peripheral organs of PD mice. Moreover, hMSCs significantly inhibited the expression of caspase-1 and NLRP3 of the NLRP3 inflammasome in both the central and peripheral organs at mRNA level. These data suggest that hMSCs protect dopaminergic neurons in PD mice by suppressing both the central and peripheral inflammatory responses, probably by inhibiting the NLRP3 inflammasome. However, the animals in our study only received several MSC infusions, and the effects of infused MSCs over extended periods on the NLRP3 inflammasome need to be investigated in future studies.

Induced neural stem cells from Macaca fascicularis show potential of dopaminergic neuron specification and efficacy in a mouse Parkinson's disease model.

Induced neural stem cells (iNSCs) can be reprogrammed from somatic cells and have shown potentials in treatment of various neurological diseases/disorders. Obtaining iNSCs of nonhuman primates serves as an important bridge for clinical translation using iNSCs. In the current study, cynomolgus (Macaca fascicularis) bone marrow mesenchymal stromal cells (MSCs) were reprogrammed into iNSCs by transduction of non-integrative Sendai virus encoding transgenes OCT4, SOX2, KLF4 and C-MYC. The obtained iNSCs showed characteristics of normal neural stem cells (NSCs) and could differentiate into neurons, astrocytes and oligodendrocytes. Furthermore, iNSCs could give rise to dopaminergic neural cells in vitro, which showed safety and efficacy after transplantation into the striatum of an immunodeficient mouse Parkinson's disease (PD) model.

The Oxidative Damage and Inflammation Mechanisms in GERD-Induced Barrett's Esophagus.

Barrett's esophagus is a major complication of gastro-esophageal reflux disease and an important precursor lesion for the development of Barrett's metaplasia and esophageal adenocarcinoma. However, the cellular and molecular mechanisms of Barrett's metaplasia remain unclear. Inflammation-associated oxidative DNA damage could contribute to Barrett's esophagus. It has been demonstrated that poly(ADP-ribose) polymerases (PARPs)-associated with ADP-ribosylation plays an important role in DNA damage and inflammatory response. A previous study indicated that there is inflammatory infiltration and oxidative DNA damage in the lower esophagus due to acid/bile reflux, and gastric acid could induce DNA damage in culture esophageal cells. This review will discuss the mechanisms of Barrett's metaplasia and adenocarcinoma underlying oxidative DNA damage in gastro-esophageal reflux disease patients based on recent clinical and basic findings.

Characterization of Human-Induced Neural Stem Cells and Derivatives following Transplantation into the Central Nervous System of a Nonhuman Primate and Rats.

Neural stem cells (NSCs) and derivatives are potential cellular sources to treat neurological diseases. In the current study, we reprogrammed human peripheral blood mononuclear cells into induced NSCs (iNSCs) and inserted GFP gene into the AAVS1 site for graft tracing. Targeted integration of GFP does not affect the proliferation and differentiation capacity of iNSCs. iNSC-GFP can be further differentiated into dopaminergic precursors (DAPs) and motor neuron precursors (MNPs), respectively. iNSCs were engrafted into the motor cortex and iNSC-DAPs into the striatum and substantia nigra (SN) of a nonhuman primate, respectively. The surviving iNSCs could respond to the microenvironment of the cortex and spontaneously differentiate into mature neurons that extended neurites. iNSC-DAPs survived well and matured into DA neurons following transplantation into the striatum and SN. iNSC-MNPs could also survive and turn into motor neurons after being engrafted into the spinal cord of rats. The results suggest that iNSCs and derivatives have a potential to be used for the treatment of neurological diseases.

The phosphorylation of the Smad2/3 linker region by nemo-like kinase regulates TGF-ß signaling.

Smad2 and Smad3 (Smad2/3) are structurally similar proteins that primarily mediate the transforming growth factor-ß (TGF-ß) signaling responsible for driving cell proliferation, differentiation, and migration. The dynamics of the Smad2/3 phosphorylation provide the key mechanism for regulating the TGF-ß signaling pathway, but the details surrounding this phosphorylation remain unclear. Here, using in vitro kinase assay coupled with mass spectrometry, we identified for the first time that nemo-like kinase (NLK) regulates TGF-ß signaling via modulation of Smad2/3 phosphorylation in the linker region. TGF-ß-mediated transcriptional and cellular responses are suppressed by NLK overexpression, whereas NLK depletion exerts opposite effects. Specifically, we discovered that NLK associates with Smad3 and phosphorylates the designated serine residues located in the linker region of Smad2 and Smad3, which inhibits phosphorylation at the C terminus, thereby decreasing the duration of TGF-ß signaling. Overall, this work demonstrates that phosphorylation on the linker region of Smad2/3 by NLK counteracts the canonical phosphorylation in response to TGF-ß signals, thus providing new insight into the mechanisms governing TGF-ß signaling transduction.

Mesenchymal Stem/Stromal Cell-Mediated Mitochondrial Transfer and the Therapeutic Potential in Treatment of Neurological Diseases.

Mesenchymal stem/stromal cells (MSCs) are multipotent stem cells that can be derived from various tissues. Due to their regenerative and immunomodulatory properties, MSCs have been extensively researched and tested for treatment of different diseases/indications. One mechanism that MSCs exert functions is through the transfer of mitochondria, a key player involved in many biological processes in health and disease. Mitochondria transfer is bidirectional and has an impact on both donor and recipient cells. In this review, we discussed how MSC-mediated mitochondrial transfer may affect cellular metabolism, survival, proliferation, and differentiation; how this process influences inflammatory processes; and what is the molecular machinery that mediates mitochondrial transfer. In the end, we summarized recent advances in preclinical research and clinical trials for the treatment of stroke and spinal cord injury, through application of MSCs and/or MSC-derived mitochondria.

Proteostasis of α-Synuclein and Its Role in the Pathogenesis of Parkinson's Disease.

Aggregation of α-Synuclein, possibly caused by disturbance of proteostasis, has been identified as a common pathological feature of Parkinson's disease (PD). However, the initiating events of aggregation have not been fully illustrated, and this knowledge may be critical to understanding the disease mechanisms of PD. Proteostasis is essential in maintaining normal cellular metabolic functions, which regulate the synthesis, folding, trafficking, and degradation of proteins. The toxicity of the aggregating proteins is dramatically influenced by its physical and physiological status. Genetic mutations may also affect the metastable phase transition of proteins. In addition, neuroinflammation, as well as lipid metabolism and its interaction with α-Synuclein, are likely to contribute to the pathogenesis of PD. In this review article, we will highlight recent progress regarding α-Synuclein proteostasis in the context of PD. We will also discuss how the phase transition status of α-Synuclein could correlate with different functional consequences in PD.

A New Image Registration Algorithm Based on Evidential Reasoning.

Image registration is a crucial and fundamental problem in image processing and computer vision, which aims to align two or more images of the same scene acquired from different views or at different times. In image registration, since different keypoints (e.g., corners) or similarity measures might lead to different registration results, the selection of keypoint detection algorithms or similarity measures would bring uncertainty. These different keypoint detectors or similarity measures have their own pros and cons and can be jointly used to expect a better registration result. In this paper, the uncertainty caused by the selection of keypoint detector or similarity measure is addressed using the theory of belief functions, and image information at different levels are jointly used to achieve a more accurate image registration. Experimental results and related analyses show that our proposed algorithm can achieve more precise image registration results compared to several prevailing algorithms.

RNF138 confers cisplatin resistance in gastric cancer cells via activating Chk1 signaling pathway.

Chemotherapy resistance represents a major issue associated with gastric cancer (GC) treatment, and arises through multiple mechanisms, including modulation of the cell-cycle check point. Several ubiquitin kinases, including RING finger protein 138 (RNF138), have been reported to mediate the G2/M phase arrest. In this study, we investigated the role of RNF138 in the development of cisplatin resistance of two GC cell lines. We show that RNF138 levels are higher in cisplatin-resistant cell lines, compared with cisplatin-sensitive cells, and RNF138 expression was elevated during drug withdrawal following the cisplatin treatment. Using gene overexpression and silencing, we analyzed the impact of altering RNF138 level on GC cell viability, apoptosis, and cell cycle phenotypes in two isogenic cisplatin-sensitive and resistant cell lines. We show that RNF138 overexpression increased GC cell viability, decreased apoptosis and delayed cell cycle progression in the cisplatin-sensitive GC cells. Conversely, RNF138 silencing produced opposite phenotypes in the cisplatin-resistant cells. Moreover, RNF138-dependent phosphorylation of Chk1 was seen in GC cells, indicating a novel connection between cisplatin-induced DNA damage and apoptosis. Collectively, these data suggest that RNF138 modulates the cisplatin resistance in the GC cells, thus serving as a potential drug target to challenge chemotherapy failure. In addition, RNF138 can also be used as a marker to monitor the development of cisplatin resistance in GC treatment.

A Clustering-Oriented Closeness Measure Based on Neighborhood Chain and Its Application in the Clustering Ensemble Framework Based on the Fusion of Different Closeness Measures.

Closeness measures are crucial to clustering methods. In most traditional clustering methods, the closeness between data points or clusters is measured by the geometric distance alone. These metrics quantify the closeness only based on the concerned data points' positions in the feature space, and they might cause problems when dealing with clustering tasks having arbitrary clusters shapes and different clusters densities. In this paper, we first propose a novel Closeness Measure between data points based on the Neighborhood Chain (CMNC). Instead of using geometric distances alone, CMNC measures the closeness between data points by quantifying the difficulty for one data point to reach another through a chain of neighbors. Furthermore, based on CMNC, we also propose a clustering ensemble framework that combines CMNC and geometric-distance-based closeness measures together in order to utilize both of their advantages. In this framework, the "bad data points" that are hard to cluster correctly are identified; then different closeness measures are applied to different types of data points to get the unified clustering results. With the fusion of different closeness measures, the framework can get not only better clustering results in complicated clustering tasks, but also higher efficiency.

Rnf138 deficiency promotes apoptosis of spermatogonia in juvenile male mice.

Spermatogenesis, the process by which haploid sperm cells are produced from a diploid precursor cell, is essential for sexual reproduction. Here, we report that RING-finger protein 138 (Rnf138) is highly expressed in testes, especially in spermatogonia and spermatocytes. The role of Rnf138 in spermatogenesis was examined using a Rnf138-knockout mouse model. Rnf138 deficiency resulted in increased apoptosis in spermatogenic cells, loss of proliferative spermatogonia, delayed development of spermatozoa and impaired fertility. The proportion of PLZF+Ki67+ cells within the PLZF+ population decreased in the knockout mice. The phenotype was further assessed by RNA-sequencing (RNA-seq), which determined that the expression levels of many genes involved in spermatogenesis were altered in the testis of Rnf138-knockout mice. Thus, Rnf138 deficiency promotes the apoptosis of spermatogenic cells, which may have been caused by the aberrant proliferation of spermatogonia in mouse testis development.

CTCF participates in DNA damage response via poly(ADP-ribosyl)ation.

CCCTC-binding factor (CTCF) plays an essential role in regulating the structure of chromatin by binding DNA strands for defining the boundary between active and heterochromatic DNA. However, the role of CTCF in DNA damage response remains elusive. Here, we show that CTCF is quickly recruited to the sites of DNA damage. The fast recruitment is mediated by the zinc finger domain and poly (ADP-ribose) (PAR). Further analyses show that only three zinc finger motifs are essential for PAR recognition. Moreover, CTCF-deficient cells are hypersensitive to genotoxic stress such as ionizing radiation (IR). Taken together, these results suggest that CTCF participate in DNA damage response via poly(ADP-ribosylation).

Increasing sensitivity to DNA damage is a potential driver for human ovarian cancer.

Ovarian cancer is one of the most common cancers among women, accounting for more deaths than any other gynecological diseases. However, the survival rate for ovarian cancer has not essentially improved over the past thirty years. Thus, to understand the molecular mechanism of ovarian tumorigenesis is important for optimizing the early diagnosis and treating this disease. In this study, we observed obvious DNA lesions, especially DNA double strand breaks (DSBs) accompanying cell cycle checkpoint activation, in the human epithelial ovarian cancer samples, which could be due to the impaired DNA response machinery. Following this line, we found that these DNA damage response-deficient primary cancer cells were hypersensitive to DNA damage and lost their ability to repair the DNA breaks, leading to genomic instability. Of note, three key DNA damage response factors, RNF8, Ku70, and FEN1 exhibited dramatically decreased expression level, implying the dysfunctional DNA repair pathways. Re-expression of wild type RNF8, Ku70, or FEN1 in these cells restored the DNA lesions and also partially rescued the cells from death. Our current study therefore proposes that accumulated DNA lesions might be a potential driver of ovarian cancer and the impaired DNA damage responders could be the targets for clinical treatment.

Ubiquitylation of Rad51d Mediated by E3 Ligase Rnf138 Promotes the Homologous Recombination Repair Pathway.

Ubiquitylation has an important role as a signal transducer that regulates protein function, subcellular localization, or stability during the DNA damage response. In this study, we show that Ring domain E3 ubiquitin ligases RNF138 is recruited to DNA damage site quickly. And the recruitment is mediated through its Zinc finger domains. We further confirm that RNF138 is phosphorylated by ATM at Ser124. However, the phosphorylation was dispensable for recruitment to the DNA damage site. Our findings also indicate that RAD51 assembly at DSB sites following irradiation is dramatically affected in RNF138-deficient cells. Hence, RNF138 is likely involved in regulating homologous recombination repair pathway. Consistently, efficiency of homologous recombination decreased observably in RNF138-depleted cells. In addition, RNF138-deficient cell is hypersensitive to DNA damage insults, such as IR and MMS. And the comet assay confirmed that RNF138 directly participated in DNA damage repair. Moreover, we find that RAD51D directly interacted with RNF138. And the recruitment of RAD51D to DNA damage site is delayed and unstable in RNF138-depleted cells. Taken together, these results suggest that RNF138 promotes the homologous recombination repair pathway.

Evidence Combination From an Evolutionary Game Theory Perspective.

Dempster-Shafer evidence theory is a primary methodology for multisource information fusion because it is good at dealing with uncertain information. This theory provides a Dempster's rule of combination to synthesize multiple evidences from various information sources. However, in some cases, counter-intuitive results may be obtained based on that combination rule. Numerous new or improved methods have been proposed to suppress these counter-intuitive results based on perspectives, such as minimizing the information loss or deviation. Inspired by evolutionary game theory, this paper considers a biological and evolutionary perspective to study the combination of evidences. An evolutionary combination rule (ECR) is proposed to help find the most biologically supported proposition in a multievidence system. Within the proposed ECR, we develop a Jaccard matrix game to formalize the interaction between propositions in evidences, and utilize the replicator dynamics to mimick the evolution of propositions. Experimental results show that the proposed ECR can effectively suppress the counter-intuitive behaviors appeared in typical paradoxes of evidence theory, compared with many existing methods. Properties of the ECR, such as solution's stability and convergence, have been mathematically proved as well.

[The research of SurgiCase CMF software in surgical simulation and prediction for mandibular asymmetry].

OBJECTIVE: To evaluate the predictive accuracy of the SurgiCase CMF software in surgical simulation and prediction for mandibular asymmetry with 3-dimensional simulation and measurement. METHODS: CBCT data of 27 patients with mandibular asymmetry were observed in CMF, and postoperative soft tissue physiognomy were predicted by simulating sagittal ramus osteotomy with or without genioplasty. The measurement parameters representing the symmetry of soft tissue were selected and the horizontal, coronal and sagittal planes were established. The results were analyzed by SPSS 19. 0. The overlap compared color grading charts were observed. RESULTS: Angles between cheilions and the horizonta plane (Ch-Ch-FH) in the simulation and postoperative soft tissues are (2. 35 ± 1. 81)° and (1. 44 ± 1. 13)°. The angles constructed among subnasale, upper lip and lower lip (Sn-UL-LL) are (4. 02 ± 3. 05)° and (2. 59 ± 1. 64)°, showing statistically different (P < 0. 01, P < 0. 05), which means that predictive accuracy of the lip canting and lip vertical deviation is relatively low. Distance between gonioi and sagittal plane (Go'-MS), distance between gonion and pogonion (Go'-Pog') and angle betweer subnasale to menton and the horizontal plane (Sn-Me'-MS) are not statistically different, which mean! high predictive accuracy of mandibular angle and chin. By observing the overlap compared color gradin-) charts, the predictive accuracy is not good in the cheek, especially in the deviate side. CONCLUSIONS: The predictive accuracy of CMF system for patients with mandibular asymmetry is relatively high, but it is not good in the lip and cheek. The software improvement is still necessary.