Exploring the immune escape mechanisms in gastric cancer patients based on the deep AI algorithms and single-cell sequencing analysis.

Gastric cancer is a prevalent and deadly malignancy, and the response to immunotherapy varies among patients. This study aimed to develop a prognostic model for gastric cancer patients and investigate immune escape mechanisms using deep machine learning and single-cell sequencing analysis. Data from public databases were analysed, and a prediction model was constructed using 101 algorithms. The high-AIDPS group, characterized by increased AIDPS expression, exhibited worse survival, genomic variations and immune cell infiltration. These patients also showed immunotherapy tolerance. Treatment strategies targeting the high-AIDPS group identified three potential drugs. Additionally, distinct cluster groups and upregulated AIDPS-associated genes were observed in gastric adenocarcinoma cell lines. Inhibition of GHRL expression suppressed cancer cell activity, inhibited M2 polarization in macrophages and reduced invasiveness. Overall, AIDPS plays a critical role in gastric cancer prognosis, genomic variations, immune cell infiltration and immunotherapy response, and targeting GHRL expression holds promise for personalized treatment. These findings contribute to improved clinical management in gastric cancer.

Effect of cognitive-behavior therapy for children with functional abdominal pain: a meta-analysis.

BACKGROUND: Cognitive-Behavior Therapy (CBT) is the validated non-pharmacological treatment for chronic pain in pediatric patients. While some suggested CBT were comparable to the usual care in reducing children's functional abdominal pain. This meta-analysis was designed to systematically review the literature for RCTs that investigated the efficacy of CBT in children with functional abdominal pain (FAP). METHODS: PubMed, Embase, and the Cochrane library were searched for papers published up to October 2022. Studies applying different CBT delivery methods (in-person, web-based, phone-based) were included in this meta-analysis to evaluate the comprehensive effectiveness of CBT compared with usual care. Weighted and standardized mean difference with the 95% confidence intervals were used for the synthesis of the results. Primary outcome was the decrease of functional disability inventory (FDI) and the secondary outcomes were the decrease of severity in pain intensity, depression, anxiety, gastrointestinal symptoms, and improvement in physical quality of life (QoL). RESULTS: A total of 10 RCTs with 1187 children were included in the final analysis. The results showed that CBT resulted in better effect in reducing functional disability inventory (SMD=-2.282, 95%CI: -4.537 to -0.027, P = 0.047), pain intensity (SMD=-0.594, 95%CI: -1.147 to -0.040, P = 0.036), and improving QoL (SMD = 14.097, 95%CI: 0.901 to 27.292, P = 0.036) compared with the control groups. Comparable effects were observed in the severity of depression (SMD=-0.493, 95%CI: -1.594 to 0.608, P = 0.380), anxiety (SMD=-0.062, 95%CI: -0.640 to 0.517, P = 0.835), and gastrointestinal symptoms (SMD=-1.096 95%CI: -2.243 to 0.050, P = 0.061) between CBT and usual treatment. CONCLUSIONS: We observed the differences in post-treatment FAP and pain intensity for children receiving CBT compared with children receiving treatment as usual. CBT in the setting of FAP demonstrates promising developments and highlights the need for future research.

ADAM33 Silencing Inhibits Vascular Smooth Muscle Cell Migration and Regulates Cytokine Secretion in Airway Vascular Remodeling via the PI3K/AKT/mTOR Pathway.

Introduction: Vascular smooth muscle cells (VSMCs) are highly involved in airway vascular remodeling in asthma. Objectives: This study aimed to investigate the mechanisms underlying the effects of a disintegrin and metalloproteinase-33 (ADAM33) gene on the migration capacity and inflammatory cytokine secretion of VSMCs. Methods: Human aortic smooth muscle cells (HASMCs) were transfected with lentiviral vectors carrying short hairpin RNA (shRNA) targeting ADAM33 or negative control vectors. The migration capacity of HASMCs was evaluated by a transwell assay. The levels of secreted inflammatory cytokines were measured using enzyme-linked immunosorbent assay (ELISA) kits. Reverse transcription-quantitative polymerase chain reaction and Western blot assays were performed to detect mRNA and protein expression levels. Results: Silencing of ADAM33 significantly inhibited the migration of HASMCs. The expression of tumor necrosis factor alpha (TNF-α) in the supernatant of HASMCs was decreased, while that of interferon gamma (IFN-γ) was increased after the transfection of shRNA targeting ADAM33. Insufficient ADAM33 expression also suppressed the expression levels of phosphatidylinositol 3-kinase (PI3K), phospho-protein kinase B (AKT), phospho-mammalian target of rapamycin (mTOR), Rho-associated protein kinases, phospho-forkhead box protein O1 (FOXO1), and cyclin D1, but it did not affect the levels of AKT, mTOR, or Rho. Conclusion: Silencing of the ADAM33 gene inhibited HASMC migration and regulated inflammatory cytokine secretion via targeting the PI3K/AKT/mTOR pathway and its downstream signaling. These data contribute to a better understanding of the regulatory mechanisms of airway vascular remodeling in asthma.

Serum Hs-CRP level and clinical significance of patients with stress ulcer caused by massive blood loss after trauma.

Stress ulcer refers to a specific type of irritation of the inner wall of the gastrointestinal tract that occurs rapidly due to acute physiological stress such as severe disease, infection, or trauma. This study investigated the serum Hs-CRP level and clinical significance of patients with stress ulcers caused by massive blood loss after trauma. For this purpose, we studied 113 patients with enormous blood loss after trauma. During the study, 26 patients developed stress ulcers. Therefore, patients with massive blood loss after trauma were divided into two groups with and without stress ulcers. In addition to clinical and demographical evaluations, serum Hs-CRP levels were measured by ELISA test method in all patients at baseline, 6, and 12 days after starting the study. Results showed that 24 patients were excluded from the study due to termination of cooperation or death. Finally, 89 patients participated in the final analysis. Of these 89 patients, 26 developed stress ulcers. There was a significant difference between the two groups with stress and non-stress ulcers in terms of mean age (P=0.001) and gender (P=0.041). Also, there was a significant difference between the two groups regarding re-bleeding (P=0.012), the number of hospitalization days (P=0.001), and a decrease in hemoglobin (P=0.035). But there was no difference between the two groups regarding the need for re-surgery (P=0.276). The results of this study showed that increased serum hs-CRP levels are directly related to stress ulcers. Patients with higher serum Hs-CRP levels were more likely to develop stress ulcers than patients without stress ulcers during six days (P=0.04) and twelve days after starting the study (P=0.001). Current research results also show that the prevalence of stress ulcers occurs in men more than women. The risk of stress ulcers increases among older patients. People with stress ulcers also lose more hemoglobin, and finally, patients with more trauma and more extended hospital stays have a higher chance of developing stress ulcers.

USP9X Increased Tumor Angiogenesis in Mantle Cell Lymphoma by Upregulation of CCND1-Mediated SOX11.

Mantle cell lymphoma (MCL) is an aggressive lymphoid malignancy with a poor prognosis. Ubiquitin-specific peptidase 9, X-linked (USP9X), has been associated with multiple physiological pathways and regulates various cellular activities. In this study, we explored the role of USP9X in MCL in vitro and in vivo. USP9X was verified to be increased in peripheral blood mononuclear cells (PBMCs) of MCL patients and MCL cells. Moreover, CCND1 and SOX11 were also upregulated in PBMCs of MCL patients. The positive correlation between USP9X and CCND1, USP9X and SOX11, and CCND1 and SOX11 were identified. Further, USP9X overexpression and knockdown were performed in MCL cells. We proved that USP9X overexpression promoted proliferation and cell cycle and suppressed cell apoptosis in MCL cells. Upregulation of angiogenesis and cell migration were induced by USP9X overexpression in MCL cells. However, the USP9X knockdown showed opposite effects. In addition, USP9X was discovered to decrease Cyclin D1 (CCND1)-mediated SOX11 expression in MCL cells. We demonstrated that SOX11 overexpression reversed USP9X knockdown-mediated angiogenesis in MCL cells. Besides, tumor formation was inhibited by USP9X knockdown in mice in vivo. In conclusion, these results revealed that USP9X promoted tumor angiogenesis in MCL via increasing CCND1-mediated SOX11.

Silencing a disintegrin and metalloproteinase­33 attenuates the proliferation of vascular smooth muscle cells via PI3K/AKT pathway: Implications in the pathogenesis of airway vascular remodeling.

Accumulating evidence suggests that pulmonary expression of a disintegrin and metalloproteinase­33 (ADAM33) serves a key role in the pathogenesis of airway remodeling­related diseases, including asthma. Airway vascular proliferation has been recognized as a key feature of airway remodeling. Our previous study showed that ADAM33 is constitutively expressed in airway vascular smooth muscle cells in patients with asthma, suggesting a potential role of ADAM33 in regulating airway vascular remodeling. Using in vitro human aortic smooth muscle cells (HASMCs) and lentiviral vector carrying short hairpin RNA for ADAM33, the present study aimed to evaluate the influence of ADAM33 silencing on the proliferation and apoptosis of HASMCs and the underlying molecular pathways. Cellular proliferation was observed using the Cell Counting Kit­8 method. Cellular apoptosis was evaluated with Annexin V­PE/7­AAD staining and flow cytometry. Reverse transcription­quantitative PCR and western blotting were used to evaluate the changes in mRNA and protein levels of involved signaling molecules. It was found that silencing of ADAM33 expression in HASMCs significantly inhibited proliferation, but induced the apoptosis of HASMCs. These changes were accompanied by inhibition of the PI3K/AKT/ERK pathway and Bcl­2, but an increase in Bax expression. These results suggested that constitutive expression of ADAM33 may be important to maintain a proliferative phenotype in HASMCs. The influences of ADAM33 on proliferation and apoptosis of HASMCs may involve regulation of PI3K/AKT/ERK and Bax/Bcl­2 pathways. These findings suggested an important role of ADAM33 in airway vascular remodeling and potential therapeutic significance of ADAM33 inhibition in airway remodeling­related diseases.

Increased expression of MCM4 is associated with poor prognosis in patients with hepatocellular carcinoma.

BACKGROUND: The minichromosome maintenance (MCM) protein complex is important for DNA replication. Moreover, the expression of specific MCM complex components has been associated with the survival of hepatocellular carcinoma (HCC) patients. However, the expression and functional roles of minichromosome maintenance complex component 4 (MCM4) in HCC development and progression have not yet been explored. We analyzed the expression and clinical significance of MCM4, including its association with liver cancer patient survival. METHODS: Oncomine, UALCAN, and HCCDB (a database of HCC expression atlas) were used to characterize the expression of MCM4 in tumor and normal tissues. The expression of MCM4 at the protein level was confirmed based on immunohistochemistry (IHC) data obtained from the Human Protein Atlas (HPA) database. The level of MCM4 was measured in tumor and adjacent normal tissues by RT-qPCR, western blot and IHC staining. The copy number alterations (CNAs) and mutations in MCM4 were analyzed by cBioPortal, whereas the co-expression genes of MCM4 in HCC were obtained from Oncomine, and used for gene ontology and pathway analysis via the NetworkAnalyst 3.0 tool, to explore the predictive signaling pathway in HCC. RESULTS: The levels of MCM4 messenger (m)RNA and protein were found to be significantly higher in liver cancer tissues than in normal liver tissues. Kaplan-Meier analysis showed that the upregulation of MCM4 was significantly negatively correlated with the survival of HCC patients. CONCLUSIONS: Our data suggest that MCM4 may be used as a potential prognostic marker and therapeutic target for HCC.

Clinical Activity and Safety of Anlotinib Combined with PD-1 Blockades for Patients with Previously Treated Small Cell Lung Cancer.

OBJECTIVE: Anlotinib was the standard monotherapy for patients with previously treated small cell lung cancer (SCLC) in recent years. Programmed cell death protein 1 (PD-1) blockade combined with antiangiogenic targeted drugs have proved to play a synergistic action for cancer treatment clinically. Consequently, the present study was to investigate the efficacy and safety of anlotinib combined with PD-1 blockades for patients with previously treated SCLC. METHODS: A total of 36 patients with SCLC who were treated with at least one previous systemic chemotherapy regimen participated in this study retrospectively. All the patients were administered with anlotinib plus PD-1 blockades therapy. Clinical activity was assessed according to the change of target lesion by imaging evidence and all the subjects were followed up regularly. Safety profiles were collected and documented during the treatment. Univariate analysis was carried out using Log rank test and multivariate analysis was adjusted by Cox regression analysis. RESULTS: All the 36 patients with previously treated SCLC were able to have their efficacy and safety profile evaluated. The best overall response of the combination regimen showed that complete response was observed in one patient, partial response was noted in 9 patients, stable disease was reported in 19 patients, progressive disease was seen in 7 patients. Therefore, the objective response rate (ORR) of the 36 patients was 27.8% (95% CI: 14.2-45.2%), disease control rate (DCR) was 80.6% (95% CI: 64.0-91.8%). Regarding the prognostic data, the median PFS and OS of the 36 patients was 4.6 months (95% CI: 3.13-6.07) and 9.3 months (95% CI: 3.30-15.30), respectively. The most common treatment-related adverse reactions were hypertension (52.8%), fatigue (47.2%), diarrhea (38.9%), hand and foot reaction (38.9%) and dermal toxicity (33.3%). Furthermore, multivariate Cox regression analysis for PFS indicated that ECOG performance status was an independent factor to predict PFS. CONCLUSION: Anlotinib combined with PD-1 blockades regimen preliminarily demonstrated encouraging efficacy and tolerable safety for patients with previously treated SCLC. The conclusion should be validated in prospective clinical trials subsequently.

Efficacy and Safety of Anlotinib for Patients with Advanced NSCLC Who Progressed After Standard Regimens and the Preliminary Analysis of an Efficacy Predictor.

BACKGROUND: The aim of this study was to investigate the efficacy and safety of anlotinib for patients with advanced non-small cell lung cancer (NSCLC) who progressed after standard regimens in real world situations and the preliminary analysis of an efficacy predictor. METHODS: A total of 118 patients with advanced NSCLC who progressed after standard regimens were included in this retrospective study. Efficacy was evaluated and toxicity profile was recorded. Progression-free survival (PFS) and overall survival (OS) were assessed using Kaplan-Meier survival curve and multivariate analysis was adjusted using Cox regression analysis. RESULTS: All of the 118 patients with NSCLC were available for evaluation of efficacy. Complete response (CR, 0 case), partial response (PR, 10 cases), stable disease (SD, 79 cases) and progressive disease (PD, 29 cases) were evaluated according to RECIST version 1.1. In consequence, objective response rate (ORR) was 8.47% and disease control rate (DCR) was 75.42%. The median PFS of the 118 patients with NSCLC was 4.3 months and the median OS was 10.3 months. The results of Cox regression analysis suggested that ECOG score was an independent factor for PFS. The toxicity profile indicated that hypertension and hand-foot syndrome were the most common adverse reactions. Additionally, the preliminary analysis of an efficacy predictor suggested that the PFS of patients with hypertension was superior to those without hypertension. CONCLUSION: Anlotinib is effective and safe for patients with advanced NSCLC who progressed after standard regimens in real world situations. Hypertension may be a biomarker for efficacy prediction.

Dynamics and controllability of droplet fusion under gas-liquid-liquid three-phase flow in a microfluidic reactor.

Gas-liquid-liquid three-phase flow systems have unique advantages of controlling reagent manipulation and improving reaction performance. However, there remains a lack of insight into the dynamics and controllability of water droplet fusion assisted by gas bubbles, particularly scaling laws for use in the design and operation of complex multiphase flow processes. In the present work, a microfluidic reactor with three T-junctions was employed to sequentially generate gas bubbles and then fuse two dispersed water droplets. The formation of the dispersed phase was divided into multiple stages, and the bubble/droplet size was correlated with operating parameters. The formation of the second dispersed droplet at the third T-junction was accompanied by the fusion of the two dispersed water droplets that were formed. It revealed a two-stage process (i.e. drainage and fusion) for the two droplets to fuse while becoming mature by breaking-up with the secondary water supply stream. In addition, a droplet contact model was employed to understand the influence on the process stability and uniformity of the merged/fused droplets by varying the surfactant concentration (in oil), the viscosity of the water phase, and the flow rates of different fluids. The study provides a deeper understanding of the droplet fusion characteristics on gas-liquid-liquid three-phase flow in microreactors for a wide range of applications.

Clinical study of water drinking test and 24-hour intraocular pressure monitoring in patients with primary open angle glaucoma.

This paper aims to observe the relevance of 24-hour intraocular pressure (IOP) monitoring and water drinking test (WDT) in patient with primary open angle glaucoma (POAG). 55 patients (90 eyes) with POAG was selected and randomly divided into operation group and drug group. Operation group, with 30 cases (48 eyes), were treat with trabeculectomy. And the other 25 cases (42 eyes) were treated with antiglaucoma medication. 24-hour intraocular pressure and WDT were measured before treatment and 6 months after treatment. The correlation between the peak value of 24-hour intraocular pressure (IOP) and intraocular pressure (IOP) fluctuation and the drinking water test were analyzed. Pearson correlation analysis showed a strong positive correlation between the peak value of diurnal IOP and the peak value of WDT IOP (r=0.758); and the Pearson correlation analysis also showed a strong positive correlation between the peak value of diurnal IOP and the peak value of WDT IOP after 6 months of POAG surgery or drug therapy (r=0.759,0.712). The peak value of IOP and IOP fluctuation in operation group were lower than those in the drug group at 6 months after operation, the difference was statistically significant. The peak value of WDT IOP can reflect the curve peak value of 24-hour IOP, which can be used to evaluate the current treatment. There were significant differences in IOP and IOP fluctuation between the drug group and the operation group at 6 months after treatment. Operation groups could achieve lower IOP and IOP fluctuation.

TIMELESS contributes to the progression of breast cancer through activation of MYC.

BACKGROUND: Breast cancer is the most common malignancy and the leading cause of cancer death among women. TIMELESS (TIM), a circadian rhythm regulator, has been recently implicated in the progression of human cancer. However, the role of TIM in the progression of breast cancer has not been well-characterized. METHODS: Immunohistochemistry (IHC) staining was used to examine TIM levels in breast cancer specimens. Mammosphere formation analysis and side population analysis were used to examine the effect of TIM on the self-renewal of breast cancer stem cells. A wound healing assay and a Transwell assay were used to determine the role of TIM in breast cancer cell migration and invasion. A soft agar growth assay in vitro and tumorigenicity in vivo were used to determine the role of TIM in tumorigenicity. RESULTS: TIM levels in both breast cancer cell lines and tissues were significantly upregulated. Patients with high TIM had poorer prognosis than patients with low TIM. Overexpression of TIM dramatically enhanced, while knockdown of TIM suppressed the self-renewal of cancer stem cells (CSCs), cell invasion and migration abilities of breast cancer cells in vitro. Moreover, overexpression of TIM significantly augmented, while knockdown of TIM reduced the tumorigenicity of breast cancer cells in vivo. Mechanism studies revealed that TIM upregulated the expression and the trans-activity of the well-known oncogene MYC. Inhibition of MYC significantly blocked the effects of TIM on CSC population, cell invasion and anchor-independent cell growth. CONCLUSION: TIM plays an important role in promoting breast cancer progression and may represent a novel therapeutic target for breast cancer.

Integrin ß4 signaling promotes mammary tumor cell adhesion to brain microvascular endothelium by inducing ErbB2-mediated secretion of VEGF.

Prior studies have indicated that the ß4 integrin promotes mammary tumor invasion and metastasis by combining with ErbB2 and amplifying its signaling capacity. However, the effector pathways and cellular functions by which the ß4 integrin exerts these effects are incompletely understood. To examine if ß4 signaling plays a role during mammary tumor cell adhesion to microvascular endothelium, we have examined ErbB2-transformed mammary tumor cells expressing either a wild-type (WT) or a signaling-defective form of ß4 (1355T). We report that WT cells adhere to brain microvascular endothelium in vitro to a significantly larger extent as compared to 1355T cells. Interestingly, integrin ß4 signaling does not exert a direct effect on adhesion to the endothelium or the underlying basement membrane. Rather, it enhances ErbB2-dependent expression of VEGF by tumor cells. VEGF in turn disrupts the tight and adherens junctions of endothelial monolayers, enabling the exposure of underlying basement membrane and increasing the adhesion of tumor cells to the intercellular junctions of endothelium. Inhibition of ErbB2 on tumor cells or the VEGFR-2 on endothelial cells suppresses mammary tumor cell adhesion to microvascular endothelium. Our results indicate that ß4 signaling regulates VEGF expression by the mammary tumor cells thereby enhancing their adhesion to microvascular endothelium.

Vascular endothelial growth factor enhances cancer cell adhesion to microvascular endothelium in vivo.

To investigate whether vascular endothelial growth factor (VEGF) enhances cancer cell adhesion to normal microvessels, we used in vivo video microscopy to measure adhesion rates of MDA-MB-435s human breast cancer cells and ErbB2-transformed mouse mammary carcinomas in the postcapillary venules of rat mesentery. An individual postcapillary venule in the mesentery was injected via a glass micropipette with cancer cells either in a perfusate of mammalian Ringer solution containing 1% bovine serum albumin as a control, or with the addition of 1 nm VEGF for test measurements. Cell adhesion was measured as either the number of adherent cells or the fluorescence intensity of adherent cells in a vessel segment for 60 min. Our results showed that during both control and VEGF treatments, the number of adherent cells increased almost linearly with time over 60 min. The VEGF treatment increased the adhesion rates of human tumour cells and mouse carcinomas 1.9-fold and 1.8-fold, respectively, over those in control conditions. We also measured cancer cell adhesion after pretreatment of cells with an antibody blocking VEGF or an antibody blocking alpha 6 integrin, and pretreatment of the microvessel with VEGF receptor (KDR/Flk-1) inhibitor, SU1498, or anti-integrin extracellular matrix ligand antibody, anti-laminin-5. All antibodies and inhibitor significantly reduced adhesion, with anti-VEGF and SU1498 reducing it the most. Our results indicate that VEGF enhances cancer cell adhesion to the normal microvessel wall, and further suggest that VEGF and its receptor, KDR/Flk-1, as well as integrins of tumour cells and their ligands at the endothelium, contribute to mammary cancer cell adhesion to vascular endothelium in vivo.

[The expression of ERCC1, DNA-PKcs protein and the relation to prognosis in non-small cell lung cancer.].

BACKGROUND: The results of many studies have proven that excision repair crosscomplementation group 1 (ERCC1) and DNA-dependent protein kinase catalytic subunit(DNA-PKcs),as the key genes in the repairment of DNA damage, are related to carcinogenesis and tumor progression. The aim of this study is to investigate the expression and clinical significance of ERCC1 and DNA-PKcs in patients with NSCLC,and analyze the relationship between their expression and the prognosis. METHODS: Paraffin-embedded operative specimens from 116 patients with non-small cell lung cancer and 12 normal lung tissues adjacent to the cancer tissues were collected, with which tissue microarray was constructed. The expression of ERCC1 and DNA-PKcs protein were detected with immunohistochemsitry. RESULTS: The positive rate of ERCC1 and DNA-PKcs expression in NSCLC was 40.5% and 75% ,respectively.There were no significant difference between ERCC1 expresson in cancer tissue and normal tissue (P =0.106).The expression of DNA-PKcs in NSCLC was significantly higher than that in normal lung tissue adjacent to the cancer tissues(Chi-Square=17.467, P =0.000). The expression of ERCC1 was closely related to differentiation of the cancer(Chi-Square=5.160, P =0.023), tumor size(Chi-Square=7.068, P =0.008) and TNM stage(Chi-Square=4.033, P =0.045), but not to age and sex of the patients, smoke, lymph node metastasis, tumors histological type. The results showed that the patients with high expression of ERCC1 seemed to have better prognosis. The five-year survival rate of NSCLC patients with ERCC1 positive expression were higher than those with ERCC1 negative expression(P =0.014).The expression of DNA-PKcs was not related to all clinical characteristics and prognosis of NSCLC patients(P >0.05). Multivariate analysis showed that TNM stage rather than ERCC1 and DNA-PKcs was the independent predictor for the prognosis for survival (P =0.000). CONCLUSIONS: The expression of ERCC1 may be associated with better survival , which may be useful to predict prognosis of NSCLC patients.

HtrA2 cleaves Apollon and induces cell death by IAP-binding motif in Apollon-deficient cells.

Apollon/BRUCE is a giant IAP protein that has BIR and UBC domains in its amino- and carboxy-terminals, respectively. Apollon binds and ubiquitylates SMAC/DIABLO and caspase9, and regulates apoptosis by facilitating proteasomal degradation of these proteins. Apollon overexpression inhibits apoptosis, while its downregulation sensitizes cells to apoptosis, suggesting that Apollon level is important for apoptosis regulation. Here we show that HtrA2/Omi catalytically cleaves Apollon with its serine protease activity. Conversely, Apollon ubiquitylates and facilitates proteasomal degradation of HtrA2 that binds to Apollon through IAP-binding motif. Thus, Apollon and HtrA2 mutually downregulate each other. Expression of catalytically active, but not inactive, HtrA2 induced apoptosis in Apollon-expressing cells. In Apollon-deficient cells, however, expression of catalytically inactive HtrA2 mutant with IAP-binding motif also induced apoptosis. These results indicate that HtrA2 induces apoptosis in two different mechanisms, one with serine protease domain and the other with IAP-binding motif, in Apollon-deficient cells.

Apollon ubiquitinates SMAC and caspase-9, and has an essential cytoprotection function.

Apollon (also known as BRUCE or BIRC6) is a large protein containing baculoviral-IAP-repeat (BIR) and ubiquitin-conjugating enzyme (UBC) domains at the amino- and carboxy termini, respectively. Apollon inhibits apoptosis, but its molecular and physiological function remains unclear. Here we report that Apollon binds to, ubiquitinates and facilitates proteasomal degradation of SMAC and caspase-9, which both contain IAP-binding motifs. Targeted disruption of Apollon in mice caused embryonic and neonatal lethality. Notably, SMAC induced apoptosis in Apollon-deficient cells, but not in Apollon-expressing cells. Furthermore, the IAP-binding motif of SMAC was required to induce apoptosis in Apollon-deficient cells. These results suggest that Apollon has an essential function in preventing SMAC-induced apoptosis.