Genetic insights into across pancreatitis types: the causal influence of immunoglobulin G N-glycosylation variants on disease risk.

Background: While a few case-control studies indicated a possible correlation of IgG N-glycosylation patterns with pancreatitis, their restricted sample sizes and methodologies prevented conclusive insights into causality or distinguishing traits across pancreatitis types. Method: We conducted a two-sample Mendelian Randomization (MR) analysis to investigate the causal relationship between 77 IgG N-glycosylation traits and various types of pancreatitis, including acute pancreatitis (AP), chronic pancreatitis (CP), alcohol acute pancreatitis (AAP), and alcohol chronic pancreatitis (ACP). This analysis utilized summary-level data from genome-wide association studies (GWAS), employing methods such as IVW, MR-Egger, and weighted median. To ensure the robustness of our findings, several sensitivity analyses, including Cochran's Q statistic, leave-one-out, MR-Egger intercept, and MR-PRESSO global test were conducted. Result: Our study uncovered the causal relationship between specific IgG N-glycosylation traits and various types of pancreatitis. Notably, an increase in genetically predicted IGP7 levels was associated with a decreased risk of developing AP. For CP, our data suggested a protective effect associated with higher levels of both IGP7 and IGP31, contrasting with increased levels of IGP27 and IGP65, which were linked to a heightened risk. Moreover, in the case of AAP, elevated IGP31 levels were causatively associated with a lower incidence, while higher IGP26 levels correlated with an increased risk for ACP. Conclusion: This study establishes causal relationship between specific IgG N-glycosylation patterns and varying risks of different pancreatitis forms, underscoring their potential as predictive biomarkers. These findings necessitate further exploration into the underlying mechanisms, promising to inform more personalized diagnostic and therapeutic strategies in pancreatitis management.

Colorectal cancer cells secreting DKK4 transform fibroblasts to promote tumour metastasis.

Wnt/ß-catenin signalling is aberrantly activated in most colorectal cancer (CRC) and is one key driver involved in the initiation and progression of CRC. However, mutations of APC gene in CRC patients retain certain activity of APC protein with decreased ß-catenin signalling and DKK4 expression significantly upregulates and represses Wnt/ß-catenin signalling in human CRC tissues, suggesting that a precisely modulated activation of the Wnt/ß-catenin pathway is essential for CRC formation and progression. The underlying reasons why a specifically reduced degree, not a fully activating degree, of ß-catenin signalling in CRC are unclear. Here, we showed that a soluble extracellular inhibitor of Wnt/ß-catenin signalling, DKK4, is an independent factor for poor outcomes in CRC patients. DKK4 secreted from CRC cells inactivates ß-catenin in fibroblasts to induce the formation of stress fibre-containing fibroblasts and myofibroblasts in culture conditions and in mouse CRC xenograft tissues, resulting in restricted expansion in tumour masses at primary sites and enhanced CRC metastasis in mouse models. Reduced ß-catenin activity by a chemical inhibitor MSAB promoted the CRC metastasis. Our findings demonstrate why reduced ß-catenin activity is needed for CRC progression and provide a mechanism by which interactions between CRC cells and stromal cells affect disease promotion.

Chloroquine Alleviates Atherosclerosis by Modulating Regulatory T Cells Through the ATM/AMPK/mTOR Signaling Pathway in ApoE -/- Mice.

BACKGROUND: Clinical observation suggests the atheroprotective effect of chloroquine and its derivatives, while its mechanism remains unclear. This study aimed to observe the protective effect of chloroquine against atherosclerosis and explore the underlying mechanism. METHODS: Ataxia telangiectasia mutated (ATM) wild-type or haploinsufficient apolipoprotein-E-knockout (ATM+/+ApoE-/- or ATM+/-ApoE-/-) mice were treated with different dosages of chloroquine. Anti-CD25 antibody was used to deplete natural Tregs in ATM+/+ApoE-/- mice. The atherosclerotic burden in different groups of mice was comprehensively evaluated by H&E staining and Masson staining. The effect of chloroquine on the regulatory T cells (Tregs) was assessed in vivo and in vitro by flow cytometry and immunohistochemical staining. The expression of related proteins was detected by real-time polymerase chain reaction and western blotting. RESULTS: In ATM+/+ApoE-/- mice, chloroquine alleviated atherosclerotic lesions, stabilized the plaque, and increased Treg counts in the atherosclerotic lesions and spleens. However, in ATM haploinsufficient mice (ATM+/-ApoE-/-), chloroquine no longer prevented atherosclerosis or impacted Treg counts. Abolishing Treg cells using an anti-CD25 antibody in vivo abrogated the atheroprotective effect of chloroquine. In vitro, chloroquine promoted the differentiation of Tregs from naïve T cells, which was accompanied by enhanced ATM/AMP-activated protein kinase (AMPK) activity and reduced downstream mammalian target of rapamycin (mTOR) activity. DISCUSSION: These findings suggest that chloroquine ameliorates atherosclerosis and stabilizes plaque by modulating Tregs differentiation through the regulation of the ATM/AMPK/mTOR pathway.

Gene expression networks involved in multiple cellular programs coexist in individual hepatocellular cancer cells.

The gene expression networks of a single cell can be used to reveal cell type- and condition-specific patterns that account for cell states, cell identity, and its responses to environmental changes. We applied single cell sequencing datasets to define mRNA patterns and visualized potential cellular capacities among hepatocellular cancer cells. The expressing numbers and levels of genes were highly heterogenous among the cancer cells. The cellular characteristics were dependent strongly on the expressing numbers and levels of genes, especially oncogenes and anti-oncogenes, in an individual cancer cell. The transcriptional activations of oncogenes and anti-oncogenes were strongly linked to inherent multiple cellular programs, some of which oppose and contend against other processes, in a cancer cell. The gene expression networks of multiple cellular programs proliferation, differentiation, apoptosis, autophagy, epithelial-mesenchymal transition, ATP production, and neurogenesis coexisted in an individual cancer cell. The findings give rise a hypothesis that a cancer cell expresses balanced combinations of genes and undergoes a given biological process by rapidly transmuting gene expressing networks.

Worldwide burden attributable to diet high in red meat from 1990 to 2019.

Introduction: Red meat overconsumption is an unhealthy behavior, while its attributed burden and epidemiological pattern remain unclear. This study aimed to describe the status and trend of how the diet high in red meat burdens the world. Material and methods: We accessed the data of summary exposure values (SEVs), deaths, and disability-adjusted life years (DALYs) with their age-standardized rates in each country from the Global Burden of Disease (GBD) Collaborative Network from 1990 to 2019. We calculated estimated annual percentage changes (EAPCs) to evaluate the trends of the disease burden. Results: The age-standardized SEV rates increased in most of the 21 GBD regions, mainly in the low-middle and middle socio-demographic index (SDI) quantiles from 1990 to 2019, while East Asia increased the most rapidly. In 2019, a diet high in red meat was responsible for 0.9 million (95% uncertainty interval (UI) 0.5 to 1.3 million) deaths and 23.9 million (95% UI 15.6 to 32.0 million) DALYs worldwide. From 1990 to 2019, the total deaths and DALYs attributable to a diet high in red meat increased by over 50%. However, the age-standardized death and DALY rates decreased by 30.3% and 23.5%, respectively, during the study period. The age-standardized death and DALY rates in the middle SDI regions surpassed those in the high SDI regions from 2002. Ischemic heart disease, diabetes mellitus, and colorectal cancer were the main causes of diet high in red meat-related deaths and DALYs. Conclusions: Increasing consumption of red meat remains a global challenge, especially in the low-middle and middle SDI countries.

Polymeric dual-modal imaging nanoprobe with two-photon aggregation-induced emission for fluorescence imaging and gadolinium-chelation for magnetic resonance imaging.

Nanoprobes that offer both fluorescence imaging (FI) and magnetic resonance imaging (MRI) can provide supplementary information and hold synergistic advantages. However, synthesis of such dual-modality imaging probes that simultaneously exhibit tunability of functional groups, high stability, great biocompatibility and desired dual-modality imaging results remains challenging. In this study, we used an amphiphilic block polymer from (ethylene glycol) methyl ether methacrylate (OEGMA) and N-(2-hydroxypropyl) methacrylamide (HPMA) derivatives as a carrier to conjugate a MR contrast agent, Gd-DOTA, and a two-photon fluorophore with an aggregation-induced emission (AIE) effect, TPBP, to construct a MR/two-photon fluorescence dual-modality contrast agent, Gd-DOTA-TPBP. Incorporation of gadolinium in the hydrophilic chain segment of the OEGMA-based carrier resulted in a high r 1 value for Gd-DOTA-TPBP, revealing a great MR imaging resolution. The contrast agent specifically accumulated in the tumor region, allowing a long enhancement duration for vascular and tumor contrast-enhanced MR imaging. Meanwhile, coupling TPBP with AIE properties to the hydrophobic chain segment of the carrier not only improved its water solubility and reduced its cytotoxicity, but also significantly enhanced its imaging performance in an aqueous phase. Gd-DOTA-TPBP was also demonstrated to act as an excellent fluorescence probe for two-photon-excited bioimaging with higher resolution and greater sensitivity than MRI. Since high-resolution, complementary MRI/FI dual-modal images were acquired at both cellular and tissue levels in tumor-bearing mice after application of Gd-DOTA-TPBP, it has great potential in the early phase of disease diagnosis.

An integrated map of fibroblastic populations in human colon mucosa and cancer tissues.

Fibroblasts and myofibroblasts are major mesenchymal cells in the lamina propria of colon mucosa and in colon cancer tissues. Detailed insight into the highly specific populations of fibroblasts and myofibroblasts is required to understand the integrity and homeostasis of human colon mucosa and colon cancer. Based on gene expression profiles of single cells, we identified fibroblast populations that produce extracellular matrix components, Wnt ligand- and BMP-secreting fibroblasts, chemokine- and chemokine ligand-generating fibroblasts, highly activated fibroblasts, immune-modulating fibroblasts, epithelial cell-modulating myofibroblasts, stimuli-responsive myofibroblasts, proliferating myofibroblasts, fibroblast-like myofibroblasts, matrix producing myofibroblasts, and contractile myofibroblasts in human colon mucosa. In colon cancer tissue, the compositions of fibroblasts and myofibroblasts were highly altered, as were the expressing patterns of genes including BMPs, Wnt ligands, chemokines, chemokine ligands, growth factors and extracellular matrix components in fibroblasts and myofibroblasts. Our work expands the working atlas of fibroblasts and myofibroblasts and provides a framework for interrogating the complexity of stromal cells in human healthy colon mucosa and colon cancer tissues.

Identification of a Novel MLPH Missense Mutation in a Chinese Griscelli Syndrome 3 Patient.

Melanophilin (MLPH) functions as a linker between RAB27A and myosin Va (MYO5A) in regulating skin pigmentation during the melanosome transport process. The MYO5A-MLPH-RAB27A ternary protein complex is required for anchoring mature melanosomes in the peripheral actin filaments of melanocytes for subsequent transfer to adjacent keratinocytes. Griscelli syndrome type 3 (GS3) is caused by mutations in the MLPH gene. So far, only five variants of MLPH associated with GS3 have been reported. Here, we reported the first patient with GS3 in a Chinese population. The proband carried a novel homozygous missense mutation (c.73G>C; p.D25H), residing in the conserved Slp homology domain of MLPH, and presented with hypopigmentation of the hair, eyebrows, and eyelashes. Light microscopy revealed the presence of abnormal pigment clumping in his hair shaft. In silico tools predicted this MLPH variant to be likely pathogenic. Using immunoblotting and immunofluorescence analysis, we demonstrated that the MLPH (D25H) variant had an inhibitory effect on melanosome transport by exhibiting perinuclear melanosome aggregation in melanocytes, and greatly reduced its binding to RAB27A, although the protein level of MLPH in the patient was not changed. Our findings suggest that MLPH (D25H) is a pathogenic variant that expands the genetic spectrum of the MLPH gene.

Prognostic value and biological function of LRRN4 in colorectal cancer.

BACKGROUND: Several nervous and nerve-related biomarkers have been detected in colorectal cancer (CRC) and can contribute to the progression of CRC. However, the role of leucine-rich repeat neuronal 4 (LRRN4), a recently identified neurogenic marker, in CRC remains unclear. METHODS: We examined the expression and clinical outcomes of LRRN4 in CRC from TCGA-COREAD mRNA-sequencing datasets and immunohistochemistry in a Chinese cohort. Furthermore, colony formation, flow cytometry, wound healing assays and mouse xenograft models were used to investigate the biological significance of LRRN4 in CRC cell lines with LRRN4 knockdown or overexpression in vitro and in vivo. In addition, weighted coexpression network analysis, DAVID and western blot analysis were used to explore the potential molecular mechanism. RESULTS: We provide the first evidence that LRRN4 expression, at both the mRNA and protein levels, was remarkably high in CRC compared to controls and positively correlated with the clinical outcome of CRC patients. Specifically, LRRN4 was an independent prognostic factor for progression-free survival and overall survival in CRC patients. Further functional experiments showed that LRRN4 promoted cell proliferation, cell DNA synthesis and cell migration and inhibited apoptosis. Knockdown of LRRN4 can correspondingly decrease these effects in vitro and can significantly suppress the growth of xenografts. Several biological functions and signaling pathways were regulated by LRRN4, including proteoglycans in cancer, glutamatergic synapse, Ras, MAPK and PI3K. LRRN4 knockdown resulted in downregulation of Akt, p-Akt, ERK1/2 and p-ERK1/2, the downstream of the Ras/MAPK signaling pathway, overexpression of LRRN4 leaded to the upregulation of these proteins. CONCLUSIONS: Our results suggest that LRRN4 could be a biological and molecular determinant to stratify CRC patients into distinct risk categories, and mechanistically, this is likely attributable to LRRN4 regulating several malignant phenotypes of neoplastic cells via RAS/MAPK signal pathways.

Dual stimuli-responsive dendronized prodrug derived from poly(oligo-(ethylene glycol) methacrylate)-based copolymers for enhanced anti-cancer therapeutic effect.

In this study, we developed an enzyme- and pH-responsive dendronized poly(oligo-(ethylene glycol) methacrylate) (pOEGMA)-doxorubicin (DOX) polymeric prodrug, which combined the pOEGMA structure with a degradable peptide dendron. The introduction of the dendron in the prodrug hindered the entanglement of brush oligo-(ethylene glycol) (OEG) chains, allowed the prodrug to possess dual stimuli-responsiveness, and mediated self-assembly of the polymeric prodrug to form stable nanoparticles (NPs). Brush conformation of polyethylene glycol (PEG) side chains endowed the NPs with long-term circulation with a half-life of 16.0 h. The dual-responsive dendritic structure enhanced cellular uptake of NPs and facilitated drug release in response to overexpressed cathepsin B and an acidic pH in the tumor microenvironment, resulting in an enhanced therapeutic effect with a tumor inhibition rate of 72.9% for 4T1 tumor-bearing mice. The NPs were demonstrated to possess great hemocompatibility and biosafety. Therefore, this strategy could provide great insight for the design of poly(oligo-(ethylene glycol) methacrylate)-based copolymers as drug delivery carriers. STATEMENT OF SIGNIFICANCE: We propose a dual-stimuli-responsive dendronized strategy for improving the cancer therapeutic effect of the poly(oligo-(ethylene glycol) methacrylate) (pOEGMA)-based drug conjugates. The introduction of the functional dendron promotes self-assembly of the polymeric prodrug into nanoparticles, hindering the entanglement of brush oligo-(ethylene glycol) (OEG) chains in the conjugated drugs. The obtained poly OEGMA-GFLG-Dendron-NH-N=DOX nanoparticles maintains long circulation, while addresses the drug release issue due to the presence of high-density PEG. The drug delivery system exhibits a high therapeutic potentcy with negligible side effects.

[Establishment of 14-Color Panel for Determining Leukocyte Subsets in Human Peripheral Blood with Flow Cytometry].

OBJECTIVE: To establish a 14-color flow cytometry protocol for the examination of leukocyte subsets in human peripheral blood. METHODS: We used cell membrane surface antibodies CD45, CD3, CD4, CD8, CD19, CD56, CD16, CD14, CD25, CD127, HLA-DR, CD123, CD11c and nucleus staining dye DAPI to establish a 14-color flow cytometry assay to determine the major cell subsets in human peripheral blood. We collected peripheral blood specimens from healthy volunteers to test for antibody titers and optimal photomultiplier tube (PMT) voltage, and to conduct single-color staining and fluorescence minus one control staining. After determining the test method and test conditions, the peripheral blood samples of 18 healthy volunteers were analyzed. RESULTS: According to the cell classification and staining index, optimal antibody mass concentrations selected were as follows: CD25 and CD127 at 8.0 µg/mL, CD45, CD3, CD14 and CD123 at 4.0 µg/mL, CD8, CD19, CD56, CD16, HLA-DR and CD11c at 2.0 µg/mL, CD4 at 1.0 µg/mL and DAPI at 0.1 µg/mL. The detection voltages for CD45, CD3, CD4, CD8, CD19, CD56, CD16, CD14, CD25, CD127, HLA-DR, CD123, CD11c and DAPI were 450 V, 410 V, 400 V, 550 V, 405 V, 500 V, 520 V, 550 V, 550 V, 400 V, 450 V, 400 V, 580 V, and 300 V, respectively. The appropriate fluorescence compensation was determined by single-color staining and fluorescence minus one controls. The 14-color flow cytometry panel was established to analyze the main subsets of leukocytes in human peripheral blood, and peripheral blood samples from 18 healthy adults were examined, obtaining the percentages of each subset of peripheral blood leukocytes and the immunophenotypes of the main subsets. CONCLUSION: We established a 14-color panel for determining leukocyte subsets in human peripheral blood by flow cytometry, which produced stable and reliable results and was easy to operate.

Integration of Flow Cytometry and Computational Analysis to Dissect the Epidermal Cellular Subsets in Keloids that Correlate with Recurrence.

Keloid disease is a benign skin disease that does not have an effective therapy. More and more research shows that epidermal abnormalities are involved in keloid pathogenesis. Little is known about the relationship between the abnormal epidermal immunophenotype and clinical outcome. Nine-color flow cytometry with computational analysis was performed to detect the altered cellular subpopulation distribution in keloid lesions. Receiver operating characteristic curves were drawn to compare predictive ability between the alteration of cell subgroup frequency and the Vancouver Scar Scale. The frequency of CD49fhi/CD29+/TLR7+ cellular subsets increased in the keloid epidermis compared with that in the healthy control. CD49fmid-hi/CD29+/TLR7+/CD24+ cellular subpopulation level was increased significantly in keloids, whereas CD49flo-mid/CD29‒/TLR7‒/CD24‒ cellular subpopulation frequency was decreased. The CD49flo/CD29‒/TLR7‒/CD24+/CD117+ cellular subpopulation showed an increased frequency during recurrence with a sensitivity of 66.7% and specificity of 91.7%. The area under the curve was 0.806 for cellular subpopulation analysis, which was higher than the area under the curve for the Vancouver Scar Scale (0.583). The alteration of keloid epidermal subpopulation frequency is related to recurrence, which will provide an optional predictive marker for keloid recurrence and a potential target subset for investigating the generation of keloid.

The first Hermansky-Pudlak syndrome type 9 patient with two novel variants in Chinese population.

Hermansky-Pudlak syndrome 9 (HPS-9) is a recessive disorder caused by BLOC1S6 gene. There are only four variants identified from four HPS-9 patients so far. Here, we reported the first HPS-9 patient in a Chinese population. He had brownish-yellow hair, white skin, brown irises with visual acuity, photophobia and nystagmus. Two novel variants, c.148G>T (p.Glu50*) and c.351dupT (p.Ile118Tyrfs*10) in BLOC1S6 gene were identified by whole-exome sequencing (WES). Absence of platelet dense granules was found by whole-mount platelet electron microscopy and Western blotting assays showed the destabilized BLOC-1 subunits. He had recurrent bruising and was found to have abnormal brain waves by electroencephalogram, but did not develop thrombopenia, immunodeficiency or other symptoms reported in other HPS-9 patients. This is the first case report of BLOC-1 mutation in a Chinese population and our findings expand the mutational spectrum of HPS genes.

Computational flow cytometric analysis to detect epidermal subpopulations in human skin.

BACKGROUND: The detection and dissection of epidermal subgroups could lead to an improved understanding of skin homeostasis and wound healing. Flow cytometric analysis provides an effective method to detect the surface markers of epidermal cells while producing high-dimensional data files. METHODS: A 9-color flow cytometric panel was optimized to reveal the heterogeneous subgroups in the epidermis of human skin. The subsets of epidermal cells were characterized using automated methods based on dimensional reduction approaches (viSNE) and clustering with Spanning-tree Progression Analysis of Density-normalized Events (SPADE). RESULTS: The manual analysis revealed differences in epidermal distribution between body sites based on a series biaxial gating starting with the expression of CD49f and CD29. The computational analysis divided the whole epidermal cell population into 25 clusters according to the surface marker phenotype with SPADE. This automatic analysis delineated the differences between body sites. The consistency of the results was confirmed with PhenoGraph. CONCLUSION: A multicolor flow cytometry panel with a streamlined computational analysis pipeline is a feasible approach to delineate the heterogeneity of the epidermis in human skin.

Imbalanced Innate Lymphoid Cells are Associated With Disease Activity and Arthritis Involvement in Patients With Systemic Lupus Erythematosus.

OBJECTIVES: This study aims to evaluate the frequency and absolute number of circulating innate lymphoid cell (ILC) subsets and their associations with clinical and serological features in systemic lupus erythematosus (SLE). PATIENTS AND METHODS: We recruited 28 SLE patients (6 males, 22 females; mean age 37.57 years; range, 18 to 56 years) and 13 healthy controls (4 males, 9 females; mean age 32.08 years; range, 19 to 48 years). Circulating ILC subsets were identified by flow cytometry. Associations between all detected cells and SLE disease activity, clinical manifestations, and serum autoantibodies were analyzed. RESULTS: In this study, significantly higher frequencies of ILC2s and ILC3s, lower frequencies of ILC1s, and higher ILC1/ILC3 and ILC1/ILC2 ratios were observed in SLE patients than in healthy controls. The frequencies and number of ILC3s were positively associated with SLE disease activity index 2000 score and anti-double stranded deoxyribonucleic acid titers in patients with SLE. Decreased ILC1 frequencies, increased ILC3 frequencies, and decreased ILC1/ILC3 and ILC2/ILC3 ratios were observed in patients with arthritis compared to those without arthritis. CONCLUSION: Our results indicated biased altered distributions of circulating ILC subsets in SLE. ILC3s were associated with SLE disease activity, and ILC1s, ILC3s, and ILC1/ILC3 and ILC2/ILC3 ratios were associated with SLE accompanied with arthritis. Taken together, these results suggest that ILCs may serve as cellular biomarkers for disease activity and arthritis involvement in SLE.

Circulating Tissue Factor-Positive Procoagulant Microparticles in Patients with Type 1 Diabetes.

AIM: To investigate the count of circulating tissue factor-positive (TF+) procoagulant microparticles (MPs) in patients with type 1 diabetes mellitus (T1DM). METHODS: This case-control study included patients with T1DM and age and sex-matched healthy volunteers. The counts of phosphatidylserine-positive (PS+) MPs and TF+PS+MPs and the subgroups derived from different cell types were measured in the peripheral blood sample of the two groups using multicolor flow cytometric assay. We compared the counts of each MP between groups as well as the ratio of the TF+PS+MPs and PS+MPs (TF+PS+MPs/PS+MPs). RESULTS: We recruited 36 patients with T1DM and 36 matched healthy controls. Compared with healthy volunteers, PS+MPs, TF+PS+MPs and TF+PS+MPs/PS+MPs were elevated in patients with T1DM (PS+MPs: 1078.5 ± 158.08 vs 686.84 ± 122.04/µL, P <0.001; TF+PS+MPs: 202.10 ± 47.47 vs 108.33 ± 29.42/µL, P <0.001; and TF+PS+MPs/PS+MPs: 0.16 ± 0.04 vs 0.19 ± 0.05, P = 0.004), mostly derived from platelet, lymphocytes and endothelial cells. In the subgroup analysis, the counts of total and platelet TF+PS+MPs were increased in patients with diabetic retinopathy (DR) and with higher HbA1c, respectively. CONCLUSION: Circulating TF+PS+MPs and those derived from platelet, lymphocytes and endothelial cells were elevated in patients with T1DM.

Targeting Myeloid-derived Suppressor Cells and Programmed Death Ligand 1 Confers Therapeutic Advantage of Ablative Hypofractionated Radiation Therapy Compared With Conventional Fractionated Radiation Therapy.

PURPOSE: Ablative hypofractionated radiation therapy (AHFRT) presents a therapeutic advantage compared with conventional fractionated radiation therapy (CFRT) for primary and oligometastatic cancers. However, the underlying mechanisms remain largely unknown. In the present study, we compared the immune alterations in response to AHFRT versus CFRT and examined the significance of immune regulations contributing to the efficacy of AHFRT. METHODS AND MATERIALS: We established subcutaneous tumors using syngeneic lung cancer and melanoma cells in both immunocompetent and immunocompromised mice and treated them with AHFRT and CFRT under the same biologically equivalent dose. RESULTS: Compared with CFRT, AHFRT significantly inhibited tumor growth in immunocompetent, but not immunocompromised, mice. On the cellular level, AHFRT reduced the recruitment of myeloid-derived suppressor cells (MDSCs) into tumors and decreased the expression of programmed death-ligand 1 (PD-L1) on those cells, which unlashed the cytotoxicity of CD8+ T cells. Through the downregulation of vascular endothelial growth factor (VEGF), AHFRT inhibited VEGF/VEGF receptor signaling, which was essential for MDSC recruitment. When combined with anti-PD-L1 antibody, AHFRT presented with greater efficacy in controlling tumor growth and improving mouse survival. By altering immune regulation, AHFRT, but not CFRT, significantly delayed the growth of secondary tumors implanted outside the irradiation field. CONCLUSIONS: Targeting MDSC recruitment and enhancing antitumor immunity are crucial for the therapeutic efficacy of AHFRT. When combined with anti-PD-L1 immunotherapy, AHFRT was more potent for cancer treatment.

Neurons generated from carcinoma stem cells support cancer progression.

Recent evidences show that nervous system acts as a crucial part of cancer microenvironment. Infiltration of nerve fibers into cancer microenvironment has an important active role in cancer progression. The stimulations of both cancer growth and metastasis by members of nervous system such as neurons and glial cells have been demonstrated. However, how the nervous system is built in cancer is largely unknown. Here we show that a fraction of cancer stem cells (CSCs) derived from patients with gastric carcinoma and colorectal carcinoma are capable of producing neurons that are involved in tumor neurogenesis and tumor growth. Cancer stem cell monoclone derived from a single cancer stem cell was able to generate neurons including sympathetic and parasympathetic neurons to take part in the nervous system in cancer tissues. Knocking down the neural cell generating capability of the human CSCs inhibited the growth of xenograft tumors in mouse model. Our data demonstrate that human CSCs are able to produce one of most important components in the cancer microenvironment that are required for cancer development and progression.

Prognostic value of CD133+ CD54+ CD44+ circulating tumor cells in colorectal cancer with liver metastasis.

In the previous study, we had showed the expression of CD133+ CD54+ CD44+ cellular subpopulation of circulating tumor cells (CTCs) was significantly associated with liver metastasis of colorectal cancer (CRC). This study aimed to explore whether this subpopulation of CTCs have a prognostic value in CRC patients. Flow cytometry was used to detect the expression of cellular subpopulations of CTCs with CD133, CD54, and CD44 in 152 CRC patients, between December 2013 and October 2014. The impact of clinicopathological factors and the expression of cellular subpopulations of CTCs on overall survival were then analyzed. CRC patients with liver metastases who underwent resection of the primary tumor accompanied by surgical treatment for metastasis had a better survival than other patients (P < 0.001). The liver metastatic CRC patients with high expression of CD133+ CD54+ (P < 0.001), CD133- CD54+ (P = 0.004), and CD133+ CD44+ CD54+ (P = 0.003) cellular subpopulations of CTCs had a worse survival than those patients with low expression. Multivariable survival analyses identified carcinoembryonic antigen levels (hazard ratio [HR] = 3.056; 95% confidence interval [CI] = 1.354-6.897; P = 0.007), treatment strategy (HR = 0.212; 95% CI = 0.056-0.808; P = 0.023), and CD133+ CD44+ CD54+ cellular subpopulation of CTCs (HR = 6.459; 95% CI = 1.461-28.558; P = 0.014) as independent prognostic factors for CRC patients with liver metastasis. CD133+ CD44+ CD54+ cellular subpopulation of CTCs has a prognostic value in CRC patients with liver metastasis, especially in the survival of CRC patients with liver metastasis who did not undergo surgical treatment for metastasis.