Region-Specific CD16+ Neutrophils Promote Colorectal Cancer Progression by Inhibiting Natural Killer Cells.

The colon is the largest compartment of the immune system, with innate immune cells exposed to antigens in the environment. However, the mechanisms by which the innate immune system is instigated are poorly defined in colorectal cancer (CRC). Here, a population of CD16+ neutrophils that specifically accumulate in CRC tumor tissues by imaging mass cytometry (IMC), immune fluorescence, and flow cytometry, which demonstrated pro-tumor activity by disturbing natural killer (NK) cells are identified. It is found that these CD16+ neutrophils possess abnormal cholesterol accumulation due to activation of the CD16/TAK1/NF-κB axis, which upregulates scavenger receptors for cholesterol intake including CD36 and LRP1. Consequently, these region-specific CD16+ neutrophils not only competitively inhibit cholesterol intake of NK cells, which interrupts NK lipid raft formation and blocks their antitumor signaling but also release neutrophil extracellular traps (NETs) to induce the death of NK cells. Furthermore, CD16-knockout reverses the pro-tumor activity of neutrophils and restored NK cell cytotoxicity. Collectively, the findings suggest that CRC region-specific CD16+ neutrophils can be a diagnostic marker and potential therapeutic target for CRC.

Granulocytes Acquire Antiapoptosis Activity and Promote Tumor Growth during Tumor Progress.

Granulocytes play important roles in cancer, and their apoptotic status is often changed by the influence of tumor environment. However, the changes and the function on granulocyte apoptosis in cancer are unclear. In this study, we used tumor-bearing mouse model and tumor patients to analyzed the apoptosis of granulocytes in different tissues by flow analysis and TUNEL fluorescence staining, and found that the percentage of apoptosis cells in granulocytes was significantly decreased in late-stage tumor-bearing mouse and patients. The in vitro co-culture experiment showed that these antiapoptotic granulocytes could significantly inhibit T cell proliferation, and RNA-seq proved that there was obvious difference on the transcriptome between these cells and control cells, particularly immune-related genes. What is important, adoptive transfer of these antiapoptotic granulocytes promoted tumor progress in mouse model. Conclusively, we found that granulocytes in late-stage tumor could delay the process of apoptosis, inhibit T cell proliferation, and acquire pro-tumor activity, which provides a new therapeutic target for tumor immunity.

Effects of intestinal trefoil factor on intestinal mucus barrier in burned mice.

Severe burns might cause intense inflammatory response and tissue ischemia and hypoxia, and these effects result in intestinal mucosal barrier damage. In this study, we evaluated the effects of recombinant human intestinal trefoil factor (rhITF) on the intestinal mucus barrier after burn injury. The results showed that rhITF could improve the intestinal mucosal damage index, decrease diamine oxidase (DAO) activity, reduce intestinal damage, and thereby alleviate intestinal mucous permeability. Severe burns were associated with subsequent decreases in the mucus thickness and the levels of hexose, and mucin, and rhITF administration might partially reverse these changes. Additional experiments showed that supplementation with rhITF markedly increased the mitochondrial respiratory control rate (RCR) and phosphorus-oxygen ratio (P/O) in intestinal tissue. Moreover, rhITF improved the intestinal mucosal blood flow (IMBF) and the levels of oxygen extraction (Oext), nitric oxide (NO) and ATP. These results suggest that ITF can improve the blood perfusion of the intestinal mucosa after severe burns, promote the transport of glutamine in the intestinal mucosa, improve the energy metabolism of goblet cells, stimulate goblet cell differentiation and maturation, promote the synthesis and secretion of intestinal mucus, and maintain the barrier function of intestinal mucus.

CD16 expression on neutrophils predicts treatment efficacy of capecitabine in colorectal cancer patients.

BACKGROUND: Early detection of capecitabine-resistance could largely increase overall survival of colorectal cancer (CRC) patients. Previous studies suggested examination of immune cells in peripheral blood would help to predict efficacy of chemotherapy. METHODS: We examined the immunological characteristics of peripheral blood in CRC patients with capecitabine treatment. We analyzed the relationships between the abnormal immune cell population in capecitabine-resistance patients and major clinical features. Furthermore, RNA sequencing, analyses of cell surface marker expression and the correlations with other major immune cell populations were performed using this population to explore the possible function of these cells. RESULTS: The expression level of CD16 on neutrophils was down-regulated in capecitabine-resistant CRC patients. Patients with CD16low/-neutrophils after capecitabine therapy had adverse clinical features. What's important, the change of CD16 expression level on neutrophils appeared much earlier than CT scan. RNA sequencing revealed that CD16low/-neutrophils in capecitabine-resistant patients had lower expression level of neutrophil-related genes, compared to CD16+neutrophils in capecitabine-sensitive patients, suggesting this CD16low/-population might be immature neutrophils. Furthermore, the expression level of CD16 on neutrophils in patients with capecitabine treatment was positively correlated with the number of anti-tumor immune cell subsets, such as CD8+T cell, CD4+T cell, NK cell and monocyte. CONCLUSIONS: Our findings indicated that CD16 expression on neutrophils in peripheral blood was a good prognostic marker for predicting efficacy of capecitabine in CRC patients.

Mechanistic study of PDIA1-catalyzed TFF3 dimerization during sepsis.

AIMS: Trefoil factor 3 (TFF3) is a gut mucosal protective molecule that is secreted by intestinal goblet cells. The dimeric structure of TFF3 enables it to function in intestinal mucosal repair and to maintain its own stability. Protein disulfide isomerase a1 (PDIA1) can directly catalyze the formation, isomerization and reduction of disulfide bonds in proteins and may play an important role in the formation of TFF3 dimer. In this study, we focused on the specific molecular mechanism of TFF3 dimerization by PDIA1 and the changes during sepsis. METHODS: We examined the changes of PDIA1 and TFF3 in sepsis rats and cell models and used a variety of experimental techniques to investigate the specific molecular mechanism of PDIA1-catalyzed TFF3 dimerization. KEY FINDINGS: We found that PDIA1 can directly catalyze the dimerization of TFF3. Our MD model proposed that two TFF3 monomers form hydrogen bonds with the region b' of PDIA1 through two stepwise reactions. Furthermore, we propose that the Cys24-Cys27 active site at the region a' of PDIA1 mediates disulfide bond formation between the Cys79 residues of each of the two TFF3 monomers via deprotonation and nucleophilic attack. During sepsis, PDIA1 is downregulated and the excessive release of nitric oxide (NO) promoted PDIA1 nitrosylation. This modification reduced PDIA1 activity, which resulted in the corresponding decrease of TFF3 dimerization and compromised TFF3 dimer function. SIGNIFICANCE: Our study revealed a novel mechanism for the inhibition of intestinal mucosal repair during sepsis and revealed novel targets for the prevention and treatment of sepsis.

The role of PDIA3 in myogenesis during muscle regeneration.

Beta 3 (ß3) integrin plays an important role in the initiation of myogenesis in adult muscle. Protein disulfide isomerases (PDIs) can activate ß3 integrin in various cells to promote cell migration, adhesion and fusion. However, the effect of PDIs on myogenesis during muscle regeneration has not been elucidated. Here, we report that PDIA3 expression is induced in regenerating myofibers. The inhibition of PDIA3 in muscle injuries in mice disrupts myoblast differentiation, impairs muscle regeneration, and ultimately aggravates muscle damage. Moreover, PDIA3 expression is upregulated and observed on the cell surfaces of myoblasts during differentiation and fusion. The inhibition of extracellular PDIA3 with an anti-PDIA3 monoclonal antibody attenuates ß3 integrin/AKT/mTOR signal activity, inhibits myoblast differentiation, and blocks the fusion of myoblasts. Thus, PDIA3 may be a mediator of myoblast differentiation and fusion during muscle regeneration.

Glutamine protects intestinal mucosa and promotes its transport after burn injury in rats.

Glutamine is an important energy source for intestinal epithelial cells (IEC); however, it is still controversial whether glutaminecan be fully utilized under pathological conditions. In this study, we investigated the changes in glutamine transport after burns and assessed the effects of exogenous glutamine administration. Finally, the potential underlying mechanisms were explored. Experimental rats were randomly divided into three groups: control group (C); burn group (B); burn+glutamine group (B+G). Rats in groups B+G and B received intragastric administration of isodose glutamine or alanine, respectively. At days 1, 3 and 5 after burns, the structure of intestinal mucosa and brush-border membrane vesicles (BBMV) were observed. The glutamine transport capacity of IEC and BBMV was detected. The synthesis of glutamine transporter ASCT2 and B0AT1 was determined. Moreover, the intestinal mucosal blood flow (IMBF), diamine oxidase activity, and the glutamine and ATP content were measured. The results showed that burn injury caused structural damage to IECs and BBMV, and significantly impaired the ability for glutamine transportation. Moreover, the mRNA and protein expressions of ASCT2 and B0AT1 as well as the glutamine and ATP content were markedly decreased. Compared with group B, most of these indicators in group B+G showed significant improvement, and approached normal levels. We conclude that glutamine administration can relieve intestinal damage, improve IMBF, promote energy synthesis and alleviate endoplasmic reticulum stress after burn injury. Finally, the synthesis and modification of ASCT2 and B0AT1 are promoted, which ultimately enhances intestinal glutamine transport.

Effects of glutamine on intestinal mucus barrier after burn injury.

Severe burns may cause intense stress and persistent inflammation, resulting in intestinal mucosal barrier damage. In this study, we evaluated the effects of glutamine (Gln) on intestinal mucus barrier after burn injury. The results showed that glutamine could improve intestinal mucosal blood flow (IMBF), decrease diamine oxidase (DAO) activity, and reduce intestine damage, thereby alleviate intestinal mucous permeability. Severe burn was associated with subsequent decrease in mucus thickness, levels of hexose, sialic acid, and protein. Glutamine administration might partially reverse these changes. Additional experiments showed that supplementation with glutamine could markedly raise the content of glutamine, glutathione (GSH), and ATP in intestinal tissue. Moreover, the levels of mRNA and protein expression of MUC2, intestinal trefoil factor (ITF) were increased remarkably, but contrary to the trend of GRP-78, CHOP. These results suggest that glutamine can improve tissue perfusion and increase energy synthesis in enterocytes, decrease endoplasmic reticulum stress (ERS) and improve mucin and ITF synthesis. Finally, lessen intestinal mucus barrier damage after burn injury.

The application of early goal directed therapy in patients during burn shock stage.

Early goal directed therapy (EGDT) provided at the earliest stages of burn shock, has significant benefits for ordinary burn patients, however, its effect on patients with more than 80% of total surface area burned (TBSA) still remains unclear. In this study, 34 extensively burned patients with (87.3±5.6)% of total surface area burned were collected from January 2008 to January 2014. All burn patients here had similar monitoring or treatment modalities. Of these 34 burn patients, 13 patients were treated with EGDT under pulse indicator continuous cardiac output (PICCO) monitoring, and 21 patients were treated with conventional fluid management. Information obtained in the course of treatment included mean arterial pressure (MAP), central venous oxygen saturation (ScvO2), oxygenation index (PaO2/FiO2), blood lactic acid and urine volume, infusion volume (mL·1% TBSA-1·Kg-1), complications of over-resuscitation (hydrothorax or pulmonary edema), case rate of burn sepsis and fatality. Our results demonstrated that there existed significant difference between the two groups in parameters below: 1. Higher ScvO2 (%) after EGDT (EGDT: 78.1±8.6, CG: 65.5±11.2; t=-3.446, P<0.05), 2. Higher PaO2/FiO2 after EGDT (EGDT: 381.4±56.6, CG: 328.9±48.6; t=2-875, P<0.05), 3. Lower mean infusion volume after EGDT (mL·1% TBSA-1·Kg-1) (EGDT: 3.29±0.26, CG: 3.71±0.31; t=5.292, P<0.05), 4. Lower complications of over-resuscitation after EGDT (EGDT: 2/13, CG: 15/21; P<0.05); However, no statistical significance existed in parameters below: 1. MAP (EGDT: 76.2±13.1, CG: 74.3±15.6; t=-0.36, P>0.05), 2. Urine volume (EGDT: 0.83±0.12, CG: 0.85±0.17; t=0.370, P>0.05), 3. Case of burn sepsis (EGDT: 13/13, CG: 20/21; P=1), 4. Case fatality (EGDT: 1/13, CG: 3/21; P=1). The finding results showed that patients with more than 80% of total surface area burned during burn shock phase could get better outcome from EGDT.