206,977 newborn screening results reveal the ethnic differences in the spectrum of inborn errors of metabolism in Huaihua, China.

Background: Inborn errors of metabolism (IEMs) are rare diseases caused by inherited defects in various biochemical pathways that strongly correlate with early neonatal mortality and stunting. Currently, no studies have reported on the incidence of IEMs of multi-ethnic groups in Huaihua, China. Methods: A total of 206,977 neonates with self-reported ethnicity who underwent IEM screening at Huaihua from 2015 to 2021 were selected for observation. Among them, 69 suspected IEM-positive neonates were referred for urine gas chromatography-mass spectrometry analysis, biochemical detection, next-generation sequencing, and Sanger sequencing. Results: Sixty-nine newborns were diagnosed with IEMs, with an overall incidence of 1:3,000. The two most common disorders were 2-methylbutyryl glycinuria (1:7,137) and phenylalanine hydroxylase deficiency (1:22,997). Moreover, the incidence of IEMs in the minority ethnic group (Miao, Dong, Tujia and Yao) (1:1,852) was markedly higher than in the Han ethnic group (1:4,741). Some ethnic features variants were identified; NM_001609.4:c.1165A>G in the ACADSB gene for Miao and Dong ethnic groups, NM_014251.2:c.852_855del in the SLC25A13 gene for Miao ethnic groups. Conclusion: This study revealed the IEM incidence within the minority ethnic groups is markedly higher than among the Han nationality and the gene variant spectrum is dramatically different in Huaihua, China. Hence, It serves as a theoretical reference for the screening and diagnosing of neonatal IEMs of multi-ethnic groups in the Huaihua area, and across China.

Microbial metabolite enhances immunotherapy efficacy by modulating T cell stemness in pan-cancer.

The immune checkpoint blockade (ICB) response in human cancers is closely linked to the gut microbiota. Here, we report that the abundance of commensal Lactobacillus johnsonii is positively correlated with the responsiveness of ICB. Supplementation with Lactobacillus johnsonii or tryptophan-derived metabolite indole-3-propionic acid (IPA) enhances the efficacy of CD8+ T cell-mediated αPD-1 immunotherapy. Mechanistically, Lactobacillus johnsonii collaborates with Clostridium sporogenes to produce IPA. IPA modulates the stemness program of CD8+ T cells and facilitates the generation of progenitor exhausted CD8+ T cells (Tpex) by increasing H3K27 acetylation at the super-enhancer region of Tcf7. IPA improves ICB responsiveness at the pan-cancer level, including melanoma, breast cancer, and colorectal cancer. Collectively, our findings identify a microbial metabolite-immune regulatory pathway and suggest a potential microbial-based adjuvant approach to improve the responsiveness of immunotherapy.

Lightweight Pneumatically Elastic Backbone Structure with Modular Construction and Nonlinear Interaction for Soft Actuators.

There has been a growing need for soft robots operating various force-sensitive tasks due to their environmental adaptability, satisfactory controllability, and nonlinear mobility unique from rigid robots. It is of desire to further study the system instability and strongly nonlinear interaction phenomenon that are the main influence factors to the actuations of lightweight soft actuators. In this study, we present a design principle on lightweight pneumatically elastic backbone structure (PEBS) with the modular construction for soft actuators, which contains a backbone printed as one piece and a common strip balloon. We build a prototype of a lightweight (<80 g) soft actuator, which can perform bending motions with satisfactory output forces (∼20 times self-weight). Experiments are conducted on the bending effects generated by interactions between the hyperelastic inner balloon and the elastic backbone. We investigated the nonlinear interaction and system instability experimentally, numerically, and parametrically. To overcome them, we further derived a theoretical nonlinear model and a numerical model. Satisfactory agreements are obtained between the numerical, theoretical, and experimental results. The accuracy of the numerical model is fully validated. Parametric studies are conducted on the backbone geometry and stiffness, balloon stiffness, thickness, and diameter. The accurate controllability, operation safety, modularization ability, and collaborative ability of the PEBS are validated by designing PEBS into a soft laryngoscope, a modularized PEBS library for a robotic arm, and a PEBS system that can operate remote surgery. The reported work provides a further applicability potential of soft robotics studies.

Heat-inactivated Bifidobacterium adolescentis ameliorates colon senescence through Paneth-like-cell-mediated stem cell activation.

Declined numbers and weakened functions of intestinal stem cells (ISCs) impair the integrity of the intestinal epithelium during aging. However, the impact of intestinal microbiota on ISCs in this process is unclear. Here, using premature aging mice (telomerase RNA component knockout, Terc-/-), natural aging mice, and in vitro colonoid models, we explore how heat-inactivated Bifidobacterium adolescentis (B. adolescentis) affects colon senescence. We find that B. adolescentis could mitigate colonic senescence-related changes by enhancing intestinal integrity and stimulating the regeneration of Lgr5+ ISCs via Wnt/ß-catenin signaling. Furthermore, we uncover the involvement of Paneth-like cells (PLCs) within the colonic stem-cell-supporting niche in the B. adolescentis-induced ISC regeneration. In addition, we identify soluble polysaccharides (SPS) as potential effective components of B. adolescentis. Overall, our findings reveal the role of heat-inactivated B. adolescentis in maintaining the ISCs regeneration and intestinal barrier, and propose a microbiota target for ameliorating colon senescence.

Bifidobacterium adolescentis orchestrates CD143+ cancer-associated fibroblasts to suppress colorectal tumorigenesis by Wnt signaling-regulated GAS1.

BACKGROUND: The interplay between gut microbiota and tumor microenvironment (TME) in the pathogenesis of colorectal cancer (CRC) is not well explored. Here, we elucidated the functional role of Bifidobacterium adolescentis (B.a) on CRC and investigated its possible mechanism on the manipulation of cancer-associated fibroblasts (CAFs) in CRC. METHODS: Different CRC animal models and various cell line models were established to explore the function of B.a on CRC. The single-cell RNA sequencing (scRNA-seq) or flow cytometry was used to detect the cell subsets in the TME of CRC. Western blot, quantitative real-time polymerase chain reaction (qRT-PCR), or immunofluorescence staining were performed to examine the activation of Wnt signaling and growth arrest specific 1 (GAS1) on CD143+ CAFs. Chromatin immunoprecipitation quantitative real-time PCR (CHIP-qPCR) was performed to investigate the regulation of transcription factor 4 (TCF4) on GAS1. Multi-immunofluorescence assay examined the expression level of CD143 and GAS1 on tissue microarray. RESULTS: We found that B.a abundance was significantly reduced in CRC patients from two independent cohorts and the bacteria database of GMrepo. Supplementation with B.a suppressed ApcMin/+ spontaneous or AOM/DSS-induced tumorigenesis in mice. scRNA-seq revealed that B.a facilitated a subset of CD143+ CAFs by inhibiting the infiltration of Th2 cells, while promoting the TNF-alpha+ B cells in TME. CD143+ CAFs highly expressed GAS1 and exhibited tumor suppressive effect. Mechanistically, GAS1 was activated by the Wnt/ß-catenin signaling in CD143+ CAFs. B.a abundance was correlated with the expression level of CD143 and GAS1. The level of CD143+ CAFs predicted the better survival outcome in CRC patients. CONCLUSIONS: These results highlighted that B.a induced a new subset of CD143+ CAFs by Wnt signaling-regulated GAS1 to suppress tumorigenesis and provided a novel therapeutic target for probiotic-based modulation of TME in CRC.

Hierarchical hollow α-Fe2O3/ZnFe2O4/Mn2O3 Janus micromotors as dynamic and efficient microcleaners for enhanced photo-Fenton elimination of organic pollutants.

Micro/nanomotors that can promote mass transport have attracted more and more research concern in the photocatalysis field. Here we first report a newly-designed hierarchical α-Fe2O3/ZnFe2O4/Mn2O3 magnetic micromotor as a heterogeneous photocatalyst for the degradation of cationic dye methylene blue (MB) from wastewater. The resulting three-dimensional (3D) flower-like hollow Janus micromotors are fabricated through a green and scalable strategy, in which each component has different functions. ZnFe2O4 microspheres serve as a magnetic scaffold for the nucleation and growth of α-Fe2O3 nanosheets and for the recycling of the micromachine. α-Fe2O3 nanosheets have shown great potential as an ideal semiconductor material for the photocatalytic decontamination of pollutants. Mn2O3 nanoparticles are mainly utilized as a catalyst to produce O2 bubbles to propel the autonomic movement of the micromotors in the presence of H2O2 fuel and also as a Fenton-like catalyst to decompose H2O2 to generate reactive oxygen species. Furthermore, the resultant micromotors exhibited linear-like motion form with an average speed of 189.1 µm s-1 in 5 wt% H2O2 solution. Moreover, the self-driven micromotors exhibited a superior catalytic degradation property toward MB, which was attributed to the synergistic effect of heterogeneous photocatalyst and the boosted micro-mixing and mass transfer caused by the vigorous motion of the micro-actuator. The possible degradation intermediates and passways of MB by α-Fe2O3/ZnFe2O4/Mn2O3 micromotor were identified with time of flight mass spectroscopy (TOF-MS). The 3D Janus micromotors have the potential to be used as a high-efficiency and active heterogeneous photocatalyst for the degradation of organic pollutants.

Bifidobacterium adolescentis induces Decorin+ macrophages via TLR2 to suppress colorectal carcinogenesis.

BACKGROUND: The interplay between gut microbiota and tumor microenvironment (TME) in the pathogenesis of colorectal cancer (CRC) is largely unknown. Here, we elucidated the functional role of B. adolescentis and its possible mechanism on the manipulation of Decorin+ macrophages in colorectal cancer. METHODS: The relative abundance of B. adolescentis in tumor or para-tumor tissue of CRC patients was analyzed. The role of B. adolescentis was explored in the CRC animal models. The single cell-RNA sequencing (scRNA-seq) was used to investigate the myeloid cells subsets in TME. The expression level of TLR2/YAP axis and its downstream Decorin in macrophages were tested by Western blot and qRT-PCR. Knockdown of Decorin in Raw264.7 was performed to investigate the effect of Decorin+ macrophages on subcutaneous tumor formation. Multi-immunofluorescence assay examined the number of Decorin+ macrophages on the CRC tissue. RESULTS: We found that the abundance of B. adolescentis was significantly reduced in tumor tissue of CRC patients. Supplementation with B. adolescentis suppressed AOM/DSS-induced tumorigenesis in mice. ScRNA-seq and animal experiment revealed that B. adolescentis increased Decorin+ macrophages. Mechanically, Decorin was activated by TLR2/YAP axis in macrophages. The abundance of B. adolescentis was correlated with the number of Decorin+ macrophages and the expression level of TLR2 in tumor tissue of CRC patients. CONCLUSIONS: These results highlight that B. adolescentis induced Decorin+ macrophages and provide a novel therapeutic target for probiotic-based modulation of immune microenvironment in CRC.

Distribution and characterization of extrachromosomal circular DNA in colorectal cancer.

Extrachromosomal circular DNA (eccDNA) has been shown to play an important role in the amplification of tumor genes and the maintenance of intra-tumor genetic heterogeneity, although its complex functional mechanism still remains to be elucidated. As the top three common malignancies in the world, colorectal cancer (CRC) has been threatening human life and health, whose tumorigenesis and development may have elusive connection with eccDNAs. Here, we described the extensive distribution of eccDNAs in the CRC tissues using Circle-seq, which range in size from hundreds to thousands of base pairs (bp). The distribution in tumor tissues had aggregation and tendency compared with random in tumor-adjacent tissues, accompanied with smaller and more regular circle lengths. After sequencing and restoring, we found that the shedding sites of eccDNAs in CRC had similar tendency in chromosome distribution, and focused on tumor-associated genes. Meanwhile, we combined RNA sequencing to explore the correlation of eccDNA differential expression in the gene transcription and signaling pathways, confirming a connection between eccDNA and RNA somewhere. Subsequently, we validated eccDNAs in CRC cell lines and the potential consistency of the junction sites of eccDNAs in CRC tissues and cell lines. Using fragments of the cationic amino acid transporter SLC7A1 to synthesize eccDNAs, we discovered the role of eccDNAs in different regions within the gene.

Fusobacterium nucleatum promotes colorectal cancer cells adhesion to endothelial cells and facilitates extravasation and metastasis by inducing ALPK1/NF-κB/ICAM1 axis.

Metastasis is the leading cause of death for colorectal cancer (CRC) patients, and the spreading tumor cells adhesion to endothelial cells is a critical step for extravasation and further distant metastasis. Previous studies have documented the important roles of gut microbiota-host interactions in the CRC malignancy, and Fusobacterium nucleatum (F. nucleatum) was reported to increase proliferation and invasive activities of CRC cells. However, the potential functions and underlying mechanisms of F. nucleatum in the interactions between CRC cells and endothelial cells and subsequent extravasation remain unclear. Here, we uncovered that F. nucleatum enhanced the adhesion of CRC cells to endothelial cells, promoted extravasation and metastasis by inducing ICAM1 expression. Mechanistically, we identified that F. nucleatum induced a new pattern recognition receptor ALPK1 to activate NF-κB pathway, resulting in the upregulation of ICAM1. Interestingly, the abundance of F. nucleatum in tumor tissues of CRC patients was positively associated with the expression levels of ALPK1 and ICAM1. Moreover, high expression of ALPK1 or ICAM1 was significantly associated with a shorter overall survival time of CRC patients. This study provides a new insight into the role of gut microbiota in engaging into the distant metastasis of CRC cells.

Akkermansia muciniphila Alleviates Dextran Sulfate Sodium (DSS)-Induced Acute Colitis by NLRP3 Activation.

Akkermansia muciniphila has been proved to play a crucial role in the progression of colitis, but its underlying mechanism remains inconclusive. In this study, we aim to investigate the effect of A. muciniphila on the development of acute colitis and explore the underlying mechanism. We found that the fecal level of A. muciniphila was decreased in ulcerative colitis (UC) patients compared to the healthy people in the GMrepo database. Oral administration of A. muciniphila strain BAA-835 significantly ameliorated the symptoms in dextran sulfate sodium (DSS)-induced acute colitis, evidenced by decreased body weight loss, colon length shortening, and colon histological inflammatory score. In addition, the number of goblet cells and the mucin family were enhanced after A. muciniphila treatment. Furthermore, proinflammatory cytokines such as tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), and monocyte chemoattractant protein 1 (MCP-1) had a downward trend. Mechanistically, the expression of NLRP3, caspase-1 p20, and IL-1ß p17 were upregulated in A. muciniphila-treated mice. Additionally, the colon tissues from high-A. muciniphila UC patients had a higher NLRP3 expression than that from low-A. muciniphila UC patients. Moreover, the upregulation of NLRP3 was observed in mouse macrophage Raw264.7 cells and bone marrow-derived macrophage (BMDM) cells after incubation with A. muciniphila. To clarify whether the protective effect of A. muciniphila in colitis depends on NLRP3, we performed the NLRP3-deficient assay in NLRP3-/- mice in vivo. The evidence showed that NLRP3 deficiency eliminated the protective effects of A. muciniphila in acute colitis. In conclusion, A. muciniphila alleviates DSS-induced acute colitis by NLRP3 activation, which enriches the mechanism and provides a new prospect for the probiotic-based treatment of colitis. IMPORTANCE The gut microbiota and host immune response interaction influences the progression of intestinal inflammatory disease. As a well-recognized next-generation probiotic, Akkermansia muciniphila has been proved to play a crucial role in the progression of colitis, but its underlying mechanism remains inconclusive. We found that oral administration of A. muciniphila strain BAA-835 significantly ameliorated the symptoms of acute colitis. Mechanistically, the expression of NLRP3 was upregulated in the A. muciniphila group, and the protective effect of A. muciniphila in colitis depends on NLRP3 activation. This enriches the mechanism and provides a new prospect for the probiotic-based treatment of colitis, which would promote a deeper understanding of the complex characteristics of A. muciniphila and provide guidance for the treatment of human colitis in the future.

Fusobacterium nucleatum promotes colorectal cancer metastasis through miR-1322/CCL20 axis and M2 polarization.

Colorectal cancer (CRC) is one of the most common malignant tumors and is associated with Fusobacterium nucleatum (F. nucleatum, Fn) infection. In this study, we explored the role of F. nucleatum in the CRC metastasis. Our results showed that the abundance of F. nucleatum was enriched in the feces and tumors of patients with CRC and tended to increase in stage IV compared to stage I in patients with metastatic CRC. Tumor-derived CCL20 activated by F. nucleatum not only increases CRC metastasis, but also participates in the reprograming of the tumor microenvironment. F. nucleatum promoted macrophage infiltration through CCL20 activation and simultaneously induced M2 macrophage polarization, enhancing the metastasis of CRC. In addition, we identified using database prediction and luciferase activity hat miR-1322, a candidate regulatory micro-RNA, could bind to CCL20 directly. F. nucleatum infection decreased the expression of miR-1322 by activating the NF-κB signaling pathway in CRC cells. In conclusion, F. nucleatum promotes CRC metastasis through the miR-1322/CCL20 axis and M2 polarization.

A. Muciniphila Suppresses Colorectal Tumorigenesis by Inducing TLR2/NLRP3-Mediated M1-Like TAMs.

The interplay between gut microbiota and the host immune system is emerging as a factor in the pathogenesis of colorectal cancer. Here, we set out to identify the effect of Akkermansia muciniphila (A. muciniphila) on colorectal cancer pathogenesis. A. muciniphila abundance was significantly reduced in patients with colorectal cancer from two independent clinical cohorts and the GMrepo dataset. Supplementation with A. muciniphila suppressed colonic tumorigenesis in ApcMin/+ mice and the growth of implanted HCT116 or CT26 tumors in nude mice. Mechanistically, A. muciniphila facilitated enrichment of M1-like macrophages in an NLRP3-dependent manner in vivo and in vitro. As a consequence, NLRP3 deficiency in macrophages attenuated the tumor-suppressive effect of A. muciniphila. In addition, we revealed that TLR2 was essential for the activation of the NF-κB/NLRP3 pathway and A. muciniphila induced M1-like macrophage response. We observed positive correlations between M1-like macrophages, NLRP3/TLR2 and A. muciniphila in patients with colorectal cancer, which corroborated these findings. In summary, A. muciniphila-induced M1-like macrophages provide a therapeutic target in the colorectal cancer tumor microenvironment.

Biodegradable scaffolds facilitate epiretinal transplantation of hiPSC-Derived retinal neurons in nonhuman primates.

Transplantation of stem cell-derived retinal neurons is a promising regenerative therapy for optic neuropathy. However, significant anatomic differences compromise its efficacy in large animal models. The present study describes the procedure and outcomes of human-induced pluripotent stem cell (hiPSC)-derived retinal sheet transplantation in primate models using biodegradable materials. Stem cell-derived retinal organoids were seeded on polylactic-coglycolic acid (PLGA) scaffolds and directed toward a retinal ganglion cell (RGC) fate. The seeded tissues showed active proliferation, typical neuronal morphology, and electrical excitability. The cellular scaffolds were then epiretinally transplanted onto the inner surface of rhesus monkey retinas. With sufficient graft-host contact provided by the scaffold, the transplanted tissues survived for up to 1 year without tumorigenesis. Histological examinations indicated survival, further maturation, and migration. Moreover, green fluorescent protein-labeled axonal projections toward the host optic nerve were observed. Cryopreserved organoids were also able to survive and migrate after transplantation. Our results suggest the potential efficacy of RGC replacement therapy in the repair of optic neuropathy for the restoration of visual function. STATEMENT OF SIGNIFICANCE: In the present study, we generated a human retinal sheet by seeding hiPSC-retinal organoid-derived RGCs on a biodegradable PLGA scaffold. We transplanted this retinal sheet onto the inner surface of the rhesus monkey retina. With scaffold support, donor cells survive, migrate and project their axons into the host optic nerve. Furthermore, an effective cryopreservation strategy for retinal organoids was developed, and the thawed organoids were also observed to survive and show cell migration after transplantation.

B. adolescentis ameliorates chronic colitis by regulating Treg/Th2 response and gut microbiota remodeling.

Inflammatory bowel disease (IBD) is defined as an immune dysregulation disease with poor prognosis. Various therapies based on gut microbe modulation have been proposed. In this study, we aim to explore the therapeutic effect of B. adolescentis on IBD, as well as the immune and microecology mechanism of B. adolescentis in IBD. The fecal level of B. adolescentis was decreased in the IBD patients compared with the normal people in our cohort and the GMrepo database. To further clarify the role of B. adolescentis in IBD, we induced chronic colitis with three cycles of dextran sulfate sodium (DSS). We found B. adolescentis gavage exhibited protective effects as evidenced by the significantly decreased diarrhea score, spleen weight, and increased colon length. Accordingly, the cumulative histological grading was decreased in the B. adolescentis administration group. In addition, tight junction protein and mucin family were enhanced after B. adolescentis treatment. Furthermore, distinct effects were found with decreased pro-inflammatory cytokines such as TNF-α, IL-6, IL-1ß, IL-18, IL-22, IL-9 and increased anti-inflammatory cytokines IL-10, IL-4, IL-5. Importantly, the colon lamina propria in the B. adolescentis group consisted of more Treg and Th2 cells, which inhibited extreme gut inflammation. Additionally, 16srRNA sequencing showed an evident increase in the B:F ratio in the B. adolescentis group. In particular, B. adolescentis application inhibited the excessive growth of Akkermansia and Escherichia-Shigella in genus level. In conclusion, B. adolescentis refined the DSS-induced chronic colitis by stimulating protective Treg/Th2 response and gut microbiota remodeling. B. adolescentis regularly treatment might improve the therapeutic effects for inflammatory bowel disease.

Evaluation of integrated modular teaching in Chinese ophthalmology trainee courses.

BACKGROUND: Before attending ophthalmology trainee courses in Zhongshan Ophthalmic Centre, the medical students from Sun Yat-sen University had finished two years of premedical education after the six-year medical courses including basic medical courses, clinical medical courses, clerkship, and research training in medical college. Integrated modular teaching using different problem-based teaching methods in ophthalmology was designed by the teaching steering committee of Zhongshan Ophthalmic Centre. This study aimed to evaluate the effectiveness and satisfaction scales of the integrated modular teaching among the trainee students. METHODS: A total of 100 medical students attending ophthalmology trainee courses in Zhongshan Ophthalmic Centre were enrolled and randomly allocated into 4 groups according to the teaching arrangement. The trainee courses consisted of several sessions delivered in multiple methods, such as "flipped classroom" session and team-based learning session. The pre- and post-class tests were delivered to evaluate the effectiveness of the integrated modular teaching. The satisfaction survey questionnaire was collected from all participants to investigate the degree of satisfaction. RESULTS: Compared with the first-day-test score, the total last-day-test score was significantly improved by a paired t-test (t = 3.288, P = 0.001). Nineteen students obtained a significant improvement in ranking increased by more than 10 in the last-day-test, whereas they failed to obtain a higher average score for daily performance than other students (t = 0.469, P = 0.654). According to the participant satisfaction questionnaires, these innovative teaching methods were considered as effective and satisfactory. CONCLUSIONS: Integrated modular teaching in ophthalmology trainee courses is effective and appreciated by the medical college students.

Islet1 and Brn3 Expression Pattern Study in Human Retina and hiPSC-Derived Retinal Organoid.

This study was conducted to determine the dynamic Islet1 and Brn3 (POU4F) expression pattern in the human fetal retina and human-induced pluripotent stem cell- (hiPSC-) derived retinal organoid. Human fetal eyes from 8 to 27 fetal weeks (Fwks), human adult retina, hiPSC-derived retinal organoid from 7 to 31 differentiation weeks (Dwks), and rhesus adult retina were collected for cyrosectioning. Immunofluorescence analysis showed that Islet1 was expressed in retinal ganglion cells in the fetal retina, human adult retina, and retinal organoids. Unexpectedly, after Fwk 20, Brn3 expression gradually decreased in the fetal retina. In the midstage of development, Islet1 was detected in bipolar and developing horizontal cells. As the photoreceptor developed, the Islet1-positive cone precursors gradually became Islet1-negative/S-opsin-positive cones. This study highlights the distinguishing characteristics of Islet1 dynamic expression in human fetal retina development and proposes more concerns which should be taken regarding Brn3 as a cell-identifying marker in mature primate retina.

Construction, expression and functional analysis of anti-B7-H4- scFv-CH3 recombinant antibody.

The B7-H4 molecule, a unique negative regulator of T lymphocytes which is overexpressed on the surface of various tumor cells, is a particularly important target candidate for tumor therapy because it can be blocked with anti-B7-H4 antibodies to inhibit the B7-H4 signaling pathway. Our previous work established an anti-B7-H4 single-chain variable fragment (scFv) library, so we have now amplified the genes encoding anti-B7-H4-scFv and human IgG1 CH3 and ligated them by overlap extension PCR to obtain a recombinant gene. After sequencing, the gene was cloned into the expression vector pET43.1a and expression was induced in E. coli BL21 (DE3) by isopropyl-ß-D-1-thiogalactopyranoside (IPTG). The protein was purified on a nickel-nitrilotriacetic acid (Ni-NTA) resin column and its antigen specificity and affinity were examined by ELISA and western blotting. We also established a Lewis lung cancer model in C57BL/6 mice to further identify the biological function of the scFv protein in vivo. The results showed that tumor volume, body weight and necrotic tissues in the control group were significantly greater than in the experimental group, indicating that selected scFvs had good biological activity and could inhibit tumor growth in tumor-bearing mice. Our work thus offers a new approach for the development of cancer-targeted therapy.

An Optimized System for Effective Derivation of Three-Dimensional Retinal Tissue via Wnt Signaling Regulation.

Effective derivation of three-dimensional (3D) retinal tissue from human-induced pluripotent stem cells (hiPSCs) could provide models for drug screening and facilitate patient-specific retinal cell replacement therapy. However, some hiPSC lines cannot undergo 3D self-organization and show inadequate differentiation efficiency to meet clinical demand. In this study, we developed an optimized system for derivation of 3D retinal tissue. We found that the Wnt signaling pathway antagonist Dickkopf-related protein 1 (DKK-1) rescued the inability of differentiated retinal progenitors to self-organize. By evaluating DKK-1 expression and supplying DKK-1 if necessary, retinal organoids were differentiated from six hiPSC lines, which were reprogramed from three common initiating cell types. Retinal tissues derived from the optimized system were well organized and capable of surviving for further maturation. Thus, using this system, we generated retinal tissues from various hiPSC lines with high efficiency. This novel system has many potential applications in regenerative therapy and precision medicine. Stem Cells 2018;36:1709-1722.