Mouse circulating extracellular vesicles contain virus-derived siRNAs active in antiviral immunity.

Induction and suppression of antiviral RNA interference (RNAi) has been observed in mammals during infection with at least seven distinct RNA viruses, including some that are pathogenic in humans. However, while the cell-autonomous immune response mediated by antiviral RNAi is gradually being recognized, little is known about systemic antiviral RNAi in mammals. Furthermore, extracellular vesicles (EVs) also function in viral signal spreading and host immunity. Here, we show that upon antiviral RNAi activation, virus-derived small-interfering RNAs (vsiRNAs) from Nodamura virus (NoV), Sindbis virus (SINV), and Zika virus (ZIKV) enter the murine bloodstream via EVs for systemic circulation. vsiRNAs in the EVs are biologically active, since they confer RNA-RNA homology-dependent antiviral activity in both cultured cells and infant mice. Moreover, we demonstrate that vaccination with a live-attenuated virus, rendered deficient in RNAi suppression, induces production of stably maintained vsiRNAs and confers protective immunity against virus infection in mice. This suggests that vaccination with live-attenuated VSR (viral suppressor of RNAi)-deficient mutant viruses could be a new strategy to induce immunity.

Efficient Dicer processing of virus-derived double-stranded RNAs and its modulation by RIG-I-like receptor LGP2.

The interferon-regulated antiviral responses are essential for the induction of both innate and adaptive immunity in mammals. Production of virus-derived small-interfering RNAs (vsiRNAs) to restrict virus infection by RNA interference (RNAi) is a recently identified mammalian immune response to several RNA viruses, which cause important human diseases such as influenza and Zika virus. However, little is known about Dicer processing of viral double-stranded RNA replicative intermediates (dsRNA-vRIs) in mammalian somatic cells. Here we show that infected somatic cells produced more influenza vsiRNAs than cellular microRNAs when both were produced by human Dicer expressed de novo, indicating that dsRNA-vRIs are not poor Dicer substrates as previously proposed according to in vitro Dicer processing of synthetic long dsRNA. We report the first evidence both for canonical vsiRNA production during wild-type Nodamura virus infection and direct vsiRNA sequestration by its RNAi suppressor protein B2 in two strains of suckling mice. Moreover, Sindbis virus (SINV) accumulation in vivo was decreased by prior production of SINV-targeting vsiRNAs triggered by infection and increased by heterologous expression of B2 in cis from SINV genome, indicating an antiviral function for the induced RNAi response. These findings reveal that unlike artificial long dsRNA, dsRNA-vRIs made during authentic infection of mature somatic cells are efficiently processed by Dicer into vsiRNAs to direct antiviral RNAi. Interestingly, Dicer processing of dsRNA-vRIs into vsiRNAs was inhibited by LGP2 (laboratory of genetics and physiology 2), which was encoded by an interferon-stimulated gene (ISG) shown recently to inhibit Dicer processing of artificial long dsRNA in cell culture. Our work thus further suggests negative modulation of antiviral RNAi by a known ISG from the interferon response.

microRNA-155-5p initiates childhood acute lymphoblastic leukemia by regulating the IRF4/CDK6/CBL axis.

Acute lymphoblastic leukemia (ALL) is a common malignancy in children. In this study, we aimed to explore putative mechanisms of microRNA-155-5p (miR-155-5p) involvement in childhood ALL (cALL) via interactions with casitas B-lineage lymphoma (CBL), interferon regulatory factor 4 (IRF4), and cyclin-dependent kinase 6 (CDK6). Bioinformatic analysis was performed initially to identify differentially expressed genes in cALL. The expression levels of miR-155-5p, CBL, IRF4, and CDK6 in peripheral blood lymphocytes from clinical ALL samples were determined using RT-qPCR and Western blot assays. A dual-luciferase reporter gene assay was used to ascertain a possible targeting relationship between miR-155-5p and CBL, CCK-8 assay and flow cytometry were used to measure cell activity and apoptosis of ALL cells. Co-IP was performed to investigate the interaction between CBL and IRF4 and the ubiquitination level of IRF4. Furthermore, in vivo validation was performed inducing xenograft tumor models with ALL cells in nude mice. As indicated by bioinformatic analysis, miR-155-5p and CDK6 were upregulated and CBL was downregulated in ALL. miR-155-5p was found to target CBL to inhibit CBL expression. miR-155-5p promoted the proliferation of ALL cells and inhibited their apoptosis by inhibiting the expression of CBL, which otherwise degraded IRF4 protein through ubiquitination, leading to inhibited CDK6 expression. Collectively, the results show that miR-155-5p can promote the development of cALL via the regulation on CBL-mediated IRF4/CDK6 axis.

Cyclophosphamide-induced GPX4 degradation triggers parthanatos by activating AIFM1.

Cyclophosphamide is an alkylating agent used to treat a variety of cancers, including leukemia. Here, we show a previously unrecognized role of cyclophosphamide in triggering the protein degradation of glutathione peroxidase 4 (GPX4), a phospholipid hydroperoxidase that protects cells from oxidative damage. Mechanistically, we found that the ubiquitin-proteasome system, but not autophagy, mediates cyclophosphamide-induced degradation of GPX4 in human leukemia cell lines. Surprisingly, cyclophosphamide-induced degradation of GPX4 leads to caspase-independent parthanatos, but not lipid peroxidation-mediated ferroptosis, through the nuclear translocation of apoptosis-inducing factor mitochondria-associated 1 (AIFM1). Consequently, the overexpression of GPX4 or the knockdown of AIFM1 limits the anticancer activity of cyclophosphamide in vitro and in xenograft tumor models. These findings establish a new framework for understanding the central role of GPX4 in blocking oxidative cell death.

lncRNA FBXL19-AS1 is a diagnosis biomarker for paediatric patients with acute myeloid leukemia.

BACKGROUND: Long non-coding RNAs (lncRNA) have emerged as novel clinical biomarkers and therapeutic targets for various tumors because of their disease- and stage-restricted expression. lncRNA FBXL19 antisense RNA 1 (FBXL19-AS1) expression has been confirmed to be up-regulated in several tumors. However, its expression and effects in paediatric acute myeloid leukemia (AML) have not been elucidated. METHODS: Serum FBXL19-AS1 expression was determined in 137 AML patients compared to 43 healthy controls ( < 0.01). RESULTS: Using receiver operating characteristic curve analysis, we observed that serum FBXL19-AS1 provided the highly diagnostic performance for the detection of AML (AUC = 0.841, < 0.001). We also examined the association between serum FBXL19-AS1 expression and clinicopathological factors, finding that its high expression was associated with French-American-British classification ( = 0.011) and cytogenetics ( = 0.021). Survival assays with the Kaplan-Meier method revealed that the overall survival ( = 0.0088) and disease-free-survival ( = 0.0027) of AML patients with high serum FBXL19-AS1 levels were distinctly shorter compared to those with low serum FBXL19-AS1 levels. More importantly, Multivariate analysis identified serum FBXL19-AS1 overexpression as an independent unfavorable prognostic factor for both overall survival and disease-free-survival of AML patients. CONCLUSIONS: Overall, our findings revealed that high expression of serum FBXL19-AS1 might be useful as a novel prognostic and diagnostic biomarker for AML patients.

In vivo near infrared fluorescence imaging and dynamic quantification of pancreatic metastatic tumors using folic acid conjugated biodegradable mesoporous silica nanoparticles.

Cancer metastasis is one of the biggest challenges in cancer treatments since it increases the likelihood that a patient will die from the disease. Therefore, the availability of techniques for the early detection and quantification of tumors is very important. We have prepared cyanine 7.5 NHS ester (Cy7.5) and folic acid (FA) conjugated biodegradable mesoporous silica nanoparticles (bMSN@Cy7.5-FA NPs) (~100 nm) for visualizing tumors in vivo. The fluorescence spectra revealed that the emission peak of bMSN@Cy7.5-FA NPs had a red-shift of 1 nm. Confocal immunofluorescent images showed that bMSN@Cy7.5-FA NPs had an excellent targeting ability for visualizing cancer cells. In vivo fluorescence imaging has been conducted using an orthotopic model for pancreatic cancer within 48 h, and the fluorescence intensity reached a maximum at a post injection time-point of 12 h, which demonstrated that the use of bMSN@Cy7.5-FA NPs provides an excellent imaging platform for tumor precision therapy in mice.

[Influence of HMGB1/MAPK/m-TOR signaling pathway on cell autophagy and chemotherapy resistance in K562 cells].

OBJECTIVE: To observe the effect of high-mobility group box 1 (HMGB1) on autophagy and chemotherapy resistance in human leukemiacell line (K562) cells, and to explore the underlying mechanisms.
 Methods: The K562 cells were cultured in vitro and divided into 6 groups: a chemotherapeutic group, a chemotherapeutic control group, a HMGB1 preconditioning group, a HMGB1 preconditioning control group, a HMGB1 siRNA group and a siRNA control group. The chemotherapeutic group was further divided into a vincristine (VCR) group, an etoposide (VP-16) group, a cytosine arabinoside (Ara-C) group, a adriamycin (ADM) group and a arsenic trioxide (As2O3) group. The cell activity was evaluated by cell counting kit-8. The protein levels of HMGB1, microtubule-associate protein1light chain3 (LC3), AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (m-TOR) were determined by Western blotting. The level of serum HMGB1 was evaluated by enzyme-linked immunosorbent assay (ELISA). The autophagy was examined by monodansylcadaverine staining and observed under transmission electron microscopy.
 Results: Compared with the control group, the cell activity was significantly decreased and the level of serum HMGB1 was significantly increased in the chemotherapeutic (VCR, VP-16, Ara-C, ADM and As2O3) groups (all P<0.05). Compared with the control group, the cell activity and the level of serum HMGB1 were significantly increased in the HMGB1 preconditioning group (both P<0.05). Compared with the siRNA control group, the cell activity and the level of serum HMGB1 were significantly decreased in the HMGB1 siRNA group (both P<0.05). Compared with the control group, the expression of LC3-II and the formation of autophagic bodies were increased in the HMGB1 preconditioning group (both P<0.05), the p-AMPK expression was increased and p-mTOR expression was decreased (both P<0.05).
 Conclusion: HMGB1 can increase the autophagy and promote chemotherapy resistance through the pathway of AMPK/m-TOR in K562 cells.

HMGB1-DNA complex-induced autophagy limits AIM2 inflammasome activation through RAGE.

High mobility group box 1 (HMGB1) is a prototype damage-associated molecular pattern (DAMP) that can induce inflammatory and immune responses alone as well as in combination with other molecules such as DNA. However, the intricate molecular mechanisms underlying HMGB1-DNA complex-mediated innate immune response remains largely elusive. In this study, we demonstrated that HMGB1-DNA complex initially induced absent in melanoma 2 (AIM2)-dependent inflammasome activation, and promoted rapid release of inflammasome-dependent early proinflammatory cytokines such as interleukin 1ß (IL-1ß). Subsequently, HMGB1-DNA complex stimulated an ATG5-dependent cellular degradation process, autophagy, which was paralleled by a cessation of AIM2 inflammasome activation and IL-1ß release. These HMGB1-DNA complex-induced inflammasome activation and autophagy were both dependent on the receptor for advanced glycation endproducts (RAGE) that recognizes a wide array of ligands (including HMGB1 and DNA). Thus, autophagy may function as a negative counter-regulatory mechanism for HMGB1-DNA complex-induced inflammasome activation, and provide a checkpoint to limit the development of inflammation.

Cancer in 22q11.2 deletion syndrome: A case report and literature review.

Clinically, the 22q11.2 deletion syndrome (22q11.2DS) is considered the most commonly detected microdeletion syndrome. Hepatoblastoma is the most prevalent malignant liver cancer in childhood. However, cases of hepatoblastoma in children with 22q11.2DS have only been reported in four patients. In this report, we present a-13-year-old male treated at our center due to growth retardation, and later diagnosed with hepatoblastoma. Whole genome sequencing (WGS) identified 22q11.2DS. Chromosomal microarray analysis (CMA) of peripheral blood sample showed a 2.9 Mb deletion of chromosome 22q11.2. While underlying mechanisms remain unclear, our literature review suggests that patients with 22q11.2DS may show an elevated risk of malignancy. After reviewing 21 previously reported cases, we identified 33 individuals with both cancer and 22q11.2 DS or DiGeorge syndrome. Of these cases, 7 out of 33 (21%) were hematologic tumors, while 26 out of 33 (78%) were solid tumors.

The mir-199b-5p encapsulated in adipocyte-derived exosomes mediates radioresistance of colorectal cancer cells by targeting JAG1.

Radiotherapy is a key treatment option for colorectal cancer, but its efficacy varies among patients. Our previous studies suggested that adipose tissue may confer the radioresistance of several abdominal tumors, such as pancreatic cancer, biliary cancer, and others. In the present work, the effects of adipocytes in regulating the radiosensitivity of colorectal cancer are explored for the first time. It was found that colony formation was increased and radiation-induced apoptosis decreased in colorectal cancer cells HCT8 and HCT116 co-cultured with adipocytes, which verified the mediation of adipocyte-driven radioresistance in colorectal cancer in vitro. Next, the colorectal cancer cells were incubated with adipocyte-derived exosomes, and a perceptible reduction in radiosensitivity was detected. Furthermore, to investigate the possible mechanisms involved, the exosomes were isolated, the encapsulated microRNAs were extracted and analyzed by small RNA sequencing. Based on bioinformatics analysis and qRT-PCR verification, miR-199b-5p was chosen for functional annotation. It was shown that miR-199b-5p expression was significantly upregulated after 6 Gy irradiation, and overexpressed miR-199b-5p significantly suppressed the radiosensitivity of HCT8 and HCT116 cells. In addition, jagged canonical Notch ligand 1(JAG1) was identified as the target gene of miR-199b-5p by using bioinformatics prediction and dual luciferase reporter gene assay. It was demonstrated that JAG1 conferred the radioresistance of colorectal cancer cells both in vivo and in vitro. Taken together, the present study demonstrates that adipocytes trigger the radioresistance of colorectal cancer cells, probably by targeting JAG1 through an adipocyte-derived exosomal miR-199b-5p.

A multicenter prospective study on the management of hepatoblastoma in children: a report from the Chinese Children's Cancer Group.

BACKGROUND: This study aimed to identify survival risk factors in Chinese children with hepatoblastoma (HB) and assess the effectiveness of the new treatment protocol proposed by the Chinese Children's Cancer Group (CCCG) in 2016. METHODS: A multicenter, prospective study that included 399 patients with HB from January 2015 to June 2020 was conducted. Patient demographics, treatment protocols, and other related information were collected. Cox regression models and Kaplan-Meier curve methods were used. RESULTS: The 4-year event-free survival (EFS) and overall survival (OS) were 76.9 and 93.5%, respectively. The 4-year EFS rates for the very-low-risk, low-risk, intermediate-risk, and high-risk groups were 100%, 91.6%, 81.7%, and 51.0%, respectively. The 4-year OS was 100%, 97.3%, 94.4%, and 86.8%, respectively. Cox regression analysis found that age, tumor rupture (R +), and extrahepatic tumor extension (E +) were independent prognostic factors. A total of 299 patients had complete remission, and 19 relapsed. Patients with declining alpha-fetoprotein (AFP) > 75% after the first two cycles of neoadjuvant chemotherapy had a better EFS and OS than those ≤ 75%. CONCLUSIONS: The survival outcome of HB children has dramatically improved since the implementation of CCCG-HB-2016 therapy. Age ≥ 8 years, R + , and E + were independent risk factors for prognosis. Patients with a declining AFP > 75% after the first two cycles of neoadjuvant chemotherapy had better EFS and OS.

Homoharringtonine-Based Induction Regimen Improved the Remission Rate and Survival Rate in Chinese Childhood AML: A Report From the CCLG-AML 2015 Protocol Study.

PURPOSE: Homoharringtonine (HHT) is commonly used for the treatment of Chinese adult AML, and all-trans retinoic acid (ATRA) has been verified in acute promyelocytic leukemia (APL). However, the efficacy and safety of HHT-based induction therapy have not been confirmed for childhood AML, and ATRA-based treatment has not been evaluated among patients with non-APL AML. PATIENTS AND METHODS: This open-label, multicenter, randomized Chinese Children's Leukemia Group-AML 2015 study was performed across 35 centers in China. Patients with newly diagnosed childhood AML were first randomly assigned to receive an HHT-based (H arm) or etoposide-based (E arm) induction regimen and then randomly allocated to receive cytarabine-based (AC arm) or ATRA-based (AT arm) maintenance therapy. The primary end points were the complete remission (CR) rate after induction therapy, and the secondary end points were the overall survival (OS) and event-free survival (EFS) at 3 years. RESULTS: We enrolled 1,258 patients, of whom 1,253 were included in the intent-to-treat analysis. The overall CR rate was significantly higher in the H arm than in the E arm (79.9% v 73.9%, P = .014). According to the intention-to-treat analysis, the 3-year OS was 69.2% (95% CI, 65.1 to 72.9) in the H arm and 62.8% (95% CI, 58.7 to 66.6) in the E arm (P = .025); the 3-year EFS was 61.1% (95% CI, 56.8 to 65.0) in the H arm and 53.4% (95% CI, 49.2 to 57.3) in the E arm (P = .022). Among the per-protocol population, who received maintenance therapy, the 3-year EFS did not differ significantly across the four arms (H + AT arm: 70.7%, 95% CI, 61.1 to 78.3; H + AC arm: 74.8%, 95% CI, 67.0 to 81.0, P = .933; E + AC arm: 72.9%, 95% CI, 65.1 to 79.2, P = .789; E + AT arm: 66.2%, 95% CI, 56.8 to 74.0, P = .336). CONCLUSION: HHT is an alternative combination regimen for childhood AML. The effects of ATRA-based maintenance are comparable with those of cytarabine-based maintenance therapy.

Arsenic Combined With All-Trans Retinoic Acid for Pediatric Acute Promyelocytic Leukemia: Report From the CCLG-APL2016 Protocol Study.

PURPOSE: Arsenic combined with all-trans retinoic acid (ATRA) is the standard of care for adult acute promyelocytic leukemia (APL). However, the safety and effectiveness of this treatment in pediatric patients with APL have not been reported on the basis of larger sample sizes. METHODS: We conducted a multicenter trial at 38 hospitals in China. Patients with newly diagnosed APL were stratified into two risk groups according to baseline WBC count and FLT3-ITD mutation. ATRA plus arsenic trioxide or oral arsenic without chemotherapy were administered to the standard-risk group, whereas ATRA, arsenic trioxide, or oral arsenic plus reduced-dose anthracycline were administered to the high-risk group. Primary end points were event-free survival and overall survival at 2 years. RESULTS: We enrolled 193 patients with APL. After a median follow-up of 28.9 months, the 2-year overall survival rate was 99% (95% CI, 97 to 100) in the standard-risk group and 95% (95% CI, 90 to 100) in the high-risk group (P = .088). The 2-year event-free survival was 97% (95% CI, 93 to 100) in the standard-risk group and 90% (95% CI, 83 to 96) in the high-risk group (P = .252). The plasma levels of arsenic were significantly elevated after treatment, with a stable effective level ranging from 42.9 to 63.2 ng/mL during treatment. In addition, plasma, urine, hair, and nail arsenic levels rapidly decreased to normal 6 months after the end of treatment. CONCLUSION: Arsenic combined with ATRA is effective and safe in pediatric patients with APL, although long-term follow-up is still needed.

Kinetic modeling and analysis of dynamic bioluminescence imaging of substrates administered by intraperitoneal injection.

BACKGROUND: Bioluminescence imaging (BLI) has been found to have diverse applications in the life sciences and medical research due to its ease of use and high sensitivity. From kinetics analysis, dynamic imaging studies have significant advantages for diagnosis when compared to traditional static imaging studies. This work focuses on modeling and quantitatively analyzing the dynamic data produced from the intraperitoneal (IP) injection of D-luciferin in longitudinal BLI, aiming to provide a powerful tool for monitoring the growth of tumors. METHODS: We constructed a three-compartment pharmacokinetic (PK) model and employed the standard Michaelis-Menten (M-M) kinetics to investigate the dynamic BLI data produced from the IP injection of D-luciferin. The 3 compartments were the plasma compartment, the non-specific compartment, and the specific compartment. The validity of this PK model was tested by the dynamic BLI data of MKN28M-luc xenograft mice, along with the published longitudinal dynamic BLI data of B16F10-luc xenograft mice. RESULTS: The R-squares between the simulated lines and the measurement were 1 and 0.99, respectively, for the mice data and the published data. In addition, the 2 kinetic macroparameters obtained reflected the rate of tumor growth in vivo. In particular, the values of macroparameters A showed a significant dependence on tumor surface area. CONCLUSIONS: The proposed PK model may be an effective tool for use in drug development programs and for monitoring the response of tumors to treatment.

The activation of antiviral RNA interference not only exists in neural progenitor cells but also in somatic cells in mammals.

The RNA interference (RNAi) pathway directs an important antiviral immunity mechanism in plants and invertebrates. Recently, we and others have demonstrated that the antiviral RNAi response is also conserved in mammals, at least to five distinct RNA viruses, including Zika virus (ZIKV). ZIKV may preferentially infect neuronal progenitor cells (NPCs) in the developing foetal brain. Ex vivo ZIKV infection induces RNAi-mediated antiviral response in human NPCs, but not in the more differentiated NPCs or somatic cells. However, litter is known about the in vivo property or function of the virus-derived small-interfering RNAs (vsiRNAs) targeting ZIKV. Here we report a surprising observation: different from ex vivo observations, viral small RNAs (vsRNAs) targeting ZIKV were produced in vivo upon infection in both central neuron system (CNS) and muscle tissues. In addition, our findings demonstrate the production of canonical vsiRNAs in murine CNS upon antiviral RNAi activation by Sindbis virus (SINV), suggesting the possibility of antiviral immune strategy applied by mammals in the CNS.

The oncogenic role of NKAP in the growth and invasion of colon cancer cells.

It has been reported that NF­κB activating protein (NKAP) is a transcriptional repressor of the Notch signaling pathway and is involved in the proliferation and survival of hematopoietic stem cells. In the present study, we aimed to investigate the effect of NKAP on the progression and metastasis of colon cancer. The results of immunohistochemical staining and western blot analysis showed that NKAP was upregulated in colon cancer tissues, and its expression was associated with colon cancer stages. CCK­8, colony formation, Transwell, and flow cytometry assays were used to demonstrate that NKAP knockdown significantly suppressed the proliferation and invasion of HCT116 and HT­29 cells, and also induced apoptosis and autophagy. By contrast, NKAP overexpression markedly promoted the proliferation and invasion of HCT­15 cells, and inhibited cell apoptosis and autophagy. Moreover, we observed that NKAP knockdown inhibited the epithelial­mesenchymal transition (EMT) process in HCT116 and HT­29 cells, while NKAP overexpression promoted EMT in HCT­15 cells. Furthermore, NKAP knockdown inhibited activation of the Akt/mTOR signaling pathway by downregulating the phosphorylation of Akt and mTOR, as well as their downstream proteins, whereas NKAP overexpression promoted the Akt/mTOR signaling pathway. Additionally, expression of P65 was downregulated by silencing of NKAP and upregulated by NKAP overexpression. These data suggest that NKAP functions as an oncogene in the growth and invasion of colon cancer in vitro.

Intravenous Administration-Oriented Pharmacokinetic Model for Dynamic Bioluminescence Imaging.

OBJECTIVE: In vivo bioluminescence imaging (BLI) is a promising tool for monitoring the growth and metastasis of tumors. However, quantitative BLI research based on intravenous (IV) injection is limited, which greatly restricts its further application. To address this problem, we designed a pharmacokinetic (PK) model which is suitable for applying on IV administration of small amounts of D-Luciferin. METHODS: After three weeks of postimplantation, mkn28-luc xenografted mice were subjected to 40-min dynamic BLI immediately following D-Luciferin intravenous injection on days 1, 3, 5, 7, and 9. Furthermore, the PK model was applied on dynamic BLI data to obtain the sum of kinetic rate constants (SKRC). RESULTS: Results showed that the SKRC values decreased rapidly with the growth of the tumor. There was a statistical difference between the SKRC values measured at different time points, while the time point of luminous intensity peak was unaffected by the growth of the tumor. CONCLUSION: In short, our results imply that dynamic BLI combined with our PK model can predict tumor growth earlier and with higher sensitivity compared to the conventional method, which is crucial for improving drug evaluation efficacy. In addition, the dynamic BLI may provide a valuable reference for the noninvasive acquiring arterial input function, which may also provide a new application prospect for hybrid PET-optical imaging.

Investigation of the influence of sampling schemes on quantitative dynamic fluorescence imaging.

Dynamic optical data from a series of sampling intervals can be used for quantitative analysis to obtain meaningful kinetic parameters of probe in vivo. The sampling schemes may affect the quantification results of dynamic fluorescence imaging. Here, we investigate the influence of different sampling schemes on the quantification of binding potential (BP) with theoretically simulated and experimentally measured data. Three groups of sampling schemes are investigated including the sampling starting point, sampling sparsity, and sampling uniformity. In the investigation of the influence of the sampling starting point, we further summarize two cases by considering the missing timing sequence between the probe injection and sampling starting time. Results show that the mean value of BP exhibits an obvious growth trend with an increase in the delay of the sampling starting point, and has a strong correlation with the sampling sparsity. The growth trend is much more obvious if throwing the missing timing sequence. The standard deviation of BP is inversely related to the sampling sparsity, and independent of the sampling uniformity and the delay of sampling starting time. Moreover, the mean value of BP obtained by uniform sampling is significantly higher than that by using the non-uniform sampling. Our results collectively suggest that a suitable sampling scheme can help compartmental modeling of dynamic fluorescence imaging provide more accurate results and simpler operations.

Improved Tumor Targeting and Longer Retention Time of NIR Fluorescent Probes Using Bioorthogonal Chemistry.

The traditional labeling method for targeted NIR fluorescence probes requires directly covalent-bonded conjugation of targeting domains and fluorophores in vitro. Although this strategy works well, it is not sufficient for detecting or treating cancers in vivo, due to steric hindrance effects that relatively large fluorophore molecules exert on the configurations and physiological functions of specific targeting domains. The copper-free, "click-chemistry"-assisted assembly of small molecules in living systems may enhance tumor accumulation of fluorescence probes by improving the binding affinities of the targeting factors. Here, we employed a vascular homing peptide, GEBP11, as a targeting factor for gastric tumors, and we demonstrate its effectiveness for in vivo imaging via click-chemistry-mediated conjugation with fluorescence molecules in tumor xenograft mouse models. This strategy showed higher binding affinities than those of the traditional conjugation method, and our results showed that the tumor accumulation of click-chemistry-mediated probes are 11-fold higher than that of directly labeled probes. The tracking life was prolonged by 12-fold, and uptake of the probes into the kidney was reduced by 6.5-fold. For lesion tumors of different sizes, click-chemistry-mediated probes can achieve sufficient signal-to-background ratios (3.5-5) for in vivo detection, and with diagnostic sensitivity approximately 3.5 times that of traditional labeling probes. The click-chemistry-assisted detection strategy utilizes the advantages of "small molecule" probes while not perturbing their physiological functions; this enables tumor detection with high sensitivity and specific selectivity.

Aberrant expression of CD133 and CD82 in patients with pediatric acute lymphoblastic leukemia and the clinical significance.

Cluster of differentiation (CD)133 is considered to be a marker of leukemia stem cells (LSCs), which are one of the primary causes of occurrence, drug resistance and relapse of acute lymphoblastic leukemia (ALL). CD82, an adhesion molecule, performs an important role in the interaction between LSCs and their niche. The purpose of the present study was to assess CD133 and CD82 expression in patients with pediatric ALL, and to evaluate the association with the clinical data. Using flow cytometric assessment and reverse transcription-polymerase chain reaction, CD133 and CD82 expression levels were measured in the bone marrow (BM) of 37 patients with newly diagnosed (ND) pediatric ALL [ALL-ND; 30 B-cell-ALL (B-ALL) and 7 T-cell-ALL (T-ALL)], in 22 patients with complete remission pediatric ALL (ALL-CR) and in 16 age-matched children without BM disease. BM plasma CD82 concentrations were measured by ELISA. The CD82 mRNA expression level in the patients with ALL-ND was significantly higher compared with that in the controls. CD82 mRNA expression levels in pediatric patients with B cell-ALL (B-ALL) were higher than those in ALL-CR patients and controls. For T-ALL, CD82 expression in ND patients was higher than in controls. CD133 mRNA expression levels in patients with pediatric B-ALL-ND were higher than that of controls and patients with ALL-CR. The frequency of CD34+ cells in pediatric ALL was significantly higher than that in controls. Frequencies of CD34+CD133+ or CD34+CD82+ cells in pediatric ALL were higher than those in controls. A positive association was observed between CD133 and CD82 mRNA expression in patients with B-ALL. A significant association was observed between CD133 mRNA expression and the hyperdiploid karyotype. Therefore, it was considered that CD133 and CD82 may serve an important role in the evolution of pediatric ALL. CD133 and CD82 should be considered as potential markers for the prognosis of patients with ALL.