MicroRNA profile as potential molecular signature for attention deficit hyperactivity disorder in children.

AIMS: Attention-deficit/hyperactivity disorder (ADHD) is a prevalent disorder of neurodevelopment in children. The diagnosis of ADHD mainly relies on the symptoms and some may be misdiagnosed due to age-based variation in behaviours. This study aimed to explore biomarkers that are greatly needed for the accurate diagnosis of ADHD. METHODS: Seven hundred and forty-two samples were retrospectively investigated in three independent cohorts, screening, training, and validation, for circulation microRNA measurement using microarray, Taqman polymerase chain reaction, and regression analysis. RESULTS: A panel of five miRNAs (miR-4516, miR-6090, miR-4763-3p, miR-4281, and miR-4466) were identified as ADHD independent risk factors that provided a high diagnostic accuracy and specificity of ADHD (AUC = 0.940 and 0.927 in the training and validation datasets, respectively). This panel of miRNAs differentiated ADHD well from control groups. After clinical improvement by treatment, the panel of miRNAs in patients and AUC changed significantly and were close to those in healthy controls. Importantly, the targets of the miRNAs identified were commonly enriched in receptor signalling pathways, ion channels, and synapse structures. CONCLUSION: Our study identified a useful panel of miRNAs that have considerable clinical value in evaluating ADHD and provide important evidence for aberrant epigenetic regulation in ADHD.

Cereblon expression is a prognostic marker in newly diagnosed POEMS syndrome treated with lenalidomide plus dexamethasone.

POEMS syndrome is a rare plasma cell disorder. Lenalidomide has recently emerged as a therapeutic option for POEMS syndrome. Cereblon has been identified as the direct target of lenalidomide, and high cereblon expression is associated with better response and outcome to lenalidomide therapy in multiple myeloma patients. Here, we analyzed the predictive value of cereblon, IKZF1, and IKZF3 in CD138+ selected plasma cells from forty-one newly diagnosed POEMS syndrome patients treated with lenalidomide in combination with dexamethasone at both gene and protein levels. We found that patients with high cereblon expression tended to achieve better hematologic response compared to those with low expression (p = 0.024 for gene expression; p = 0.01 for protein expression). Multivariate Cox regression analysis revealed high cereblon mRNA expression as an independent prognostic marker for longer progression-free survival (hazard ratio 0.542; 95% CI 0.337-0.871; p = 0.011). In conclusion, our results emphasized the role of cereblon mRNA expression as a unique biomarker for predicting the clinical response and outcome of lenalidomide-based therapy in newly diagnosed POEMS syndrome patients.

A prospective phase II study of low dose lenalidomide plus dexamethasone in patients with newly diagnosed polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy, and skin changes (POEMS) syndrome.

Polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy, and skin changes (POEMS) syndrome is a rare plasma dyscrasia without standard treatment. This phase II prospective trial evaluates the safety and response of 12 cycles of low dose lenalidomide (10 mg) plus dexamethasone (Rdex) in patients with newly diagnosed POEMS syndrome. Forty-one patients (28 men) were enrolled and the median age at diagnosis was 49 years (range, 21-70 years). Twenty-one patients (46%) achieved complete hematologic response and the neurologic response rate was 95%. The median serum vascular endothelial growth factor (VEGF) declined from 5155 pg/mL (range, 534-14 328 pg/mL) to 832 pg/mL (95-6254 pg/mL) after therapy. The overall VEGF response rate was 83%, and the median time to response was 2 months, with a mean VEGF reduction of 43% at the first month. In terms of clinical response, Rdex substantially relieved extravascular volume overload, organomegaly, and pulmonary hypertension. No treatment-related deaths occurred and no patients suffered from lenalidomide-related grade 3 or above adverse events. After a median follow-up of 34 months, median overall survival (OS) and progression-free survival (PFS) were not reached, with an estimated 3-year OS and PFS of 90% and 75%, respectively. In conclusion, Rdex was active with high hematologic, VEGF and organ response rate and well tolerated for patients with newly diagnosed POEMS syndrome. This trial was registered at www.clinicaltrials.gov as #NCT01816620.

Vimentin is important in the neural differentiation of PC12 cells promoted by sialylation.

Sialic acid modification is a kind of post-translational modification. To investigate the regulation effect of sialic acid on neural differentiation, we used CycloManN propanyl perac (CycloManN pro), a metabolic precursor of sialic acid, to treat PC12 cells. We noted that CycloManN pro indeed robustly promoted global sialylation detected by MAL II lectin blot in PC12 cells. Simultaneously, we interestingly found that the neurite outgrowth of PC12 cells was significantly promoted by the CycloManN pro treatment. The profile analysis of sialylated proteins showed that a protein band at 55KD was greatly enhanced especially in PC12L cells after CycloManN pro treatment. After enrichment with lectin MAL II, the proteins in this band were analyzed by mass spectrometry. The results showed that 23 proteins were in the band, but the score of vimentin was the highest among them. To investigate further the role of vimentin in the process of neurite differentiation, vimentin construct was transfected into PC12 cells. We interestingly observed that ectopic expression of vimentin significantly enhanced the neurite outgrowth induced by CycloManN pro. However, after three potential glycosylation sites (Ser-7, Thr-33, Ser-34:) of vimentin were mutated to alanine, overexpression of the mutated vimentin completely lost the enhancement activity for the neural differentiation even in the presence of CycloManN pro. Taken together, our study demonstrated that vimentin was important in the induction of neural differentiation by CycloManN pro.

OCULAR MANIFESTATIONS AND TREATMENT OUTCOMES IN CHINESE PATIENTS WITH POEMS SYNDROME.

PURPOSE: To evaluate the relationship of serum vascular endothelial growth factor (VEGF) levels and ocular manifestations in Chinese patients with POEMS syndrome. METHODS: This is a prospective study. Forty-one treatment-naive patients were enrolled from April 2014 to November 2014. Among the 41 patients, 40 had complete ocular examination, spectral domain optical coherence tomography scan, and serum VEGF measurement before treatment and every 3-month interval after lenalidomide and dexamethasone treatment. RESULTS: Twenty-seven (67.5%) patients had optic disk edema (ODE) at baseline. Retinal manifestations included retinal hemorrhage, subretinal fluid, macular edema, and cotton wool spot. The difference in mean serum VEGF concentrations between patients with and without ODE was significant (P = 0.017). Among patients with ODE, there was a significant positive correlation between mean serum VEGF levels and the binocular mean retinal nerve fiber layer thickness (P = 0.008), as well as mean peripapillary retinal thickness (P = 0.020) before treatment. After 3 months to 17 months treatment, mean serum VEGF concentrations decreased significantly (P < 0.001). Mean retinal nerve fiber layer thickness and mean peripapillary retinal thickness decreased significantly (P < 0.001). The remission rate of ODE was 87.5%, and complete remission rate was 58.3%. CONCLUSION: The ODE is a common manifestation in POEMS syndrome, and raised VEGF might explain the development and mechanism. Systemic treatment could lead to decrease in serum VEGF levels accompanied by regression of ODE.

Renal impairment in patients with polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy and skin changes syndrome: incidence, treatment and outcome.

BACKGROUND: Polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy and skin changes (POEMS) syndrome is a multisystem disorder arising from underlying plasma cell dyscrasia. Renal impairment and related pathological changes have been reported, but data on its prevalence, response to therapy and impact on survival are still lacking. METHODS: We retrospectively reviewed 299 patients diagnosed with POEMS syndrome in a tertiary-care university hospital from 2000 until 2014. The estimated glomerular filtration rate (eGFR) was used to define renal impairment and response, according to International Myeloma Working Group criteria. We examined the impact of renal impairment and response on patient survival. RESULTS: Sixty-seven patients (22.4%) had renal impairment (eGFR < 60 mL/min/1.73 m(2)) at baseline. In a multivariate analysis, ascites was independently associated with renal impairment [odds ratio (OR) 12.366, P < 0.001]. Renal impairment was reversible in 66.0% of patients receiving therapy and was associated with a shorter time interval between symptom onset and treatment (OR 0.059, P = 0.043) and a vascular endothelial growth factor remission (OR 15.958, P = 0.050) in a multivariate analysis. In terms of therapy, patients with a renal response more commonly received a novel agent-based regimen (P = 0.037), which also led to a shorter response time (P = 0.001). With a median follow-up of 27.4 months, inferior survival was observed in patients with severe renal impairment (eGFR < 30 mL/min/1.73 m(2)), but not in those with moderate dysfunction (eGFR 30-59 mL/min/1.73 m(2)), compared with patients without renal impairment. A renal response, if achieved, predicted improved survival. CONCLUSIONS: Renal impairment is a common complication of POEMS syndrome, but can be reversed with effective therapy in most cases.

Prognostic value of serum heavy/light chain ratios in patients with POEMS syndrome.

POEMS syndrome is a rare plasma cell dyscrasia. Serum concentrations of the monoclonal protein in this disorder are typically low, and inapplicable to monitor disease activity in most cases, resulting in limited practical and prognostic values. Novel immunoassays measuring isotype-specific heavy/light chain (HLC) pairs showed its utility in disease monitoring and outcome prediction in several plasma cell dyscrasias. We report results of HLC measurements in 90 patients with POEMS syndrome. Sixty-six patients (73%; 95% confidence interval, 63-82%) had an abnormal HLC ratio at baseline. It could stratify the risk of disease relapse and was strongly associated with worse progression-free survival in a multivariate analysis (P = 0.021; hazard ratio [HR] 6.89, 95% CI 1.34-35.43). After therapy, HLC ratios improved, with 43 patients (48%) remaining abnormal. The post-therapeutic HLC ratio, if abnormal, also remained as an independent prognostic factor associated with worse progression-free survival (P = 0.019; HR 4.30, 95% CI 1.27-14.56). These results suggest the prognostic utility of HLC ratios in clinical management of POEMS patients.

Efficacy and safety of low-dose lenalidomide plus dexamethasone in patients with relapsed or refractory POEMS syndrome.

Although autologous stem cell transplantation or melphalan-based chemotherapy has significantly improved the prognosis of POEMS syndrome, a few patients will relapse or be refractory to primary therapy, and there is a lack of studies regarding these patients. In this study, we used low-dose lenalidomide (10 mg daily) and dexamethasone (40 mg, once weekly) to treat twelve patients with relapsed (n = 8) or refractory (n = 4) POEMS syndrome. After a median follow-up time of 20 months, the overall hematologic response rate was 77% with 44% having a complete response. Eight (67%) patients had neurological response, and the median overall neuropathy limitation scale score was reduced from 3 (range, 1-9) to 2 (range, 0-6). Serum vascular endothelial growth factor response rate was 91% and 46% of patients had normal serum VEGF levels. One patient had progression of the disease 3 months after the end of treatment and subsequently died from the disease. Therefore, the estimated 2 year overall survival and progression-free survival were 92%. The low-dose lenalidomide and dexamethasone regimen was well tolerated, with no treatment-related death or any grade 3 or 4 toxicity. In conclusion, low-dose lenalidomide plus dexamethasone therapy is an effective and safe regimen for patients with relapsed or refractory POEMS syndrome.

Sulfatide epigenetically regulates miR-223 and promotes the migration of human hepatocellular carcinoma cells.

BACKGROUND & AIMS: The biological relevance and regulation mechanism of aberrant miR-223 expression in human hepatocellular carcinoma (HCC) remain unknown. Our aim was to investigate miR-223 regulation in HCC. METHODS: miR-223 and integrin αV dysregulation were verified in 57 HCC specimens. Immunohistochemical analysis of integrin αV and sulfatide levels was performed on another cohort of 103 HCC samples. Epigenetic analysis was used to explore the effect of sulfatide on miR-223 transcription. Orthotopic growth, and intrahepatic and pulmonary metastasis of tumors derived from SMMC-7721 cells expressing miR-223 or cerebroside sulfotransferase were monitored in mice. RESULTS: miR-223 was reduced in HCC specimens and highly metastatic cell lines. Enhanced miR-223 expression had a negative effect on integrin αV-mediated cell migration. In vivo assays of metastasis in an orthotopically implanted model demonstrated that miR-223 effectively inhibited HCC metastasis. Further analysis demonstrated that integrin αV is negatively regulated by miR-223. Moreover, the integrin αV subunit was significantly positively correlated with highly expressed sulfatide in 103 HCC specimens. Intriguingly, miR-223 expression was suppressed by sulfatide in HCC in association with reduced recruitment of acetylated histone H3 and C/EBPα to the pre-miR-223 gene promoter, where monocytic leukemia zinc finger (MOZ) protein, a MYST-type histone acetyltransferase, lost its attachment. The expression of histone deacetylases, HDAC9 and HDAC10, were greatly stimulated by sulfatide and their recruitment to miR-223 gene promoter was enhanced. CONCLUSIONS: Downregulation of miR-223 in HCC is associated with the epigenetic regulation by highly expressed sulfatide and involved in tumor metastasis.

Regulation of integrin αV subunit expression by sulfatide in hepatocellular carcinoma cells.

Integrin is important in migration and metastasis of tumor cells. Changes of integrin expression and distribution will cause an alteration of cellular adhesion and migration behaviors. In this study, we investigated sulfatide regulation of the integrin αV subunit expression in hepatoma cells and observed that either exogenous or endogenous sulfatide elicited a robust upregulation of integrin αV subunit mRNA and protein expression in hepatoma cells. This regulatory effect occurred with a corresponding phosphorylation (T739) of the transcription factor Sp1. Based on the electrophoretic mobility shift assay, sulfatide enhanced the integrin αV promoter activity and strengthened the Sp1 complex super-shift. The results of chromatin immunoprecipitation analysis also indicated that sulfatide enhanced Sp1 binding to the integrin αV promoter in vivo. Silence of Sp1 diminished the stimulation of integrin αV expression by sulfatide. In the early stage of sulfatide stimulation, phosphorylation of Erk as well as c-Src was noted, and inhibition of Erk activation with either U0126 or PD98059 significantly suppressed Sp1 phosphorylation and integrin αV expression. We demonstrated that sulfatide regulated integrin αV expression and cell adhesion, which was associated with Erk activation.

Prognostic and immunological role of sulfatide-related lncRNAs in hepatocellular carcinoma.

Background: Hepatocellular carcinoma (HCC) is the most common primary malignancy of the liver. Long non-coding RNAs (lncRNAs) play important roles in the occurrence and development of HCC through multiple pathways. Our previous study reported the specific molecular mechanism for sulfatide regulation of integrin αV expression and cell adhesion in HCC cells through lncRNA AY927503. Next, it is necessary to identify more sulfatide-related lncRNAs, explore their clinical signifcance, and determine new targeted treatment strategies. Methods: Microarrays were used to screen a complete set of lncRNAs with different expression profiles in sulfatide-treated cells. Sulfatide-related lncRNAs expression data and corresponding HCC patient survival information were obtained from the The Cancer Genome Atlas (TCGA) database, and the prognosis prediction model was constructed based on Cox regression analysis. Methylated RNA immunoprecipitation with next generation sequencing (MeRIP-seq) was used to detemine the effect of sulfatide on lncRNAs m6A modification. Tumor Immune Estimation Resource (TIMER) and Gene set nnrichment analysis (GSEA) were utilized to enrich the immune and functional pathways of sulfatide-related lncRNAs. Results: A total of 85 differentially expressed lncRNAs (|Fold Change (FC)|>2, P<0.05) were screened in sulfatide-treated HCC cells. As a result, 24 sulfatide-related lncRNAs were highly expressed in HCC tissues, six of which were associated with poor prognosis in HCC patients. Based on thses data, a sulfatide-related lncRNAs prognosis assessment model for HCC was constructed. According to this risk score analysis, the overall survival (OS) curve showed that the OS of high-risk patients was significantly lower than that of low-risk patients (P<0.05). Notably, the expression difference in sulfatide-related lncRNA NRSN2-AS1 may be related to sulfatide-induced RNA m6A methylation. In addition, the expression level of NRSN2-AS1 was significantly positively correlated with immune cell infiltration in HCC and participated in the peroxisome and Peroxisome proliferator-activated receptor (PPAR) signaling pathways. Conclusions: In conclusion, sulfatide-related lncRNAs might be promising prognostic and therapeutic targets for HCC.

LncRNA BC promotes lung adenocarcinoma progression by modulating IMPAD1 alternative splicing.

BACKGROUND: The therapeutic value of targeted therapies in patients with lung cancer is reduced when tumours acquire secondary resistance after an initial period of successful treatment. However, the molecular events behind the resistance to targeted therapies in lung cancer remain largely unknown. AIMS: To discover the important role and mechanism of lncRNA BC in promoting tumor metastasis and influencing clinical prognosis of LUAD. MATERIALS & METHODS: Microarrays were used to screen a comprehensive set of lncRNAs with differential expression profiles in lung cancer cells. The functional role and mechanism of lncRNA were further investigated by gain- and loss-of-function assays. RNA pull-down, protein assays, and mass spectrometry were used to identify proteins that interacted with lncRNA. TaqMan PCR was used to measure lncRNA in lung adenocarcinoma and adjacent nontumor tissues from 428 patients. The clinical significance of lncRNA identified was statistically confirmed in this cohort of patients. RESULTS: In this study, we show that the long non-coding RNA BC009639 (BC) is involved in acquired resistance to EGFR-targeted therapies. Among the 235 long non-coding RNAs that were differentially expressed in lung cancer cell lines, with different metastatic potentials, BC promoted growth, invasion, metastasis, and resistance to EGFR-tyrosine kinase inhibitors (EGFR-TKIs), both in vitro and in vivo. BC was highly expressed in 428 patients with lung adenocarcinoma (LUAD) and high BC expression correlated with reduced efficacy of EGFR-TKI therapy. To uncover the molecular mechanism of BC-mediated EGFR-TKI resistance in lung cancer, we screened and identified nucleolin and hnRNPK that interact with BC. BC formed the splicing complex with nucleolin and hnRNPK to facilitate the production of a non-protein-coding inositol monophosphatase domain containing 1 (IMPAD1) splice variant, instead of the protein-coding variant. The BC-mediated alternative splicing (AS) of IMPAD1 resulted in the induction of the epithelial-mesenchymal transition and resistance to EGFR-TKI in lung cancer. High BC expression correlated with clinical progress and poor survival among 402 patients with LUAD. DISSCUSSION: Through alternative splicing, BC boosted the non-coding IMPAD1-203 transcript variant while suppressing the IMPAD1-201 variant. In order to control the processing of pre-mRNA, BC not only attracted RNA binding proteins (NCL, IGF2BP1) or splicing factors (hnRNPK), but also controlled the formation of the splicing-regulator complex by creating RNA-RNA-duplexes. CONCLUSION: Our results reveal an important role for BC in mediating resistance to EGFR-targeted therapy in LUAD through IMPAD1 AS and in implication for the targeted therapy resistance.

LncRNA lncAY is upregulated by sulfatide via Myb/MEF2C acetylation to promote the tumorigenicity of hepatocellular carcinoma cells.

LncRNA (long noncoding RNA) is often dysregulated in tumors especially hepatocellular carcinoma (HCC). However, the dysregulation mechanism of lncRNAs is largely unknown. Here, we showed that lncRNA lncAY expression was stimulated in HCC by either endogenous or exogenous sulfatide. Elevated lncAY promoted HCC cell migration or angiogenesis, whereas lncAY silence suppressed HCC cell migration and proliferation. Interestingly, the activity of lncAY gene promoter was enhanced by sulfatide. Then Myb and MEF2C were identified as the transcription factors responsible for the stimulation of lncAY promoter activity and transcription by sulfatide. Both Myb and MEF2C enrichment on lncAY promoter was further confirmed, and their occupancy on lncAY promoter was strengthened by sulfatide for Myb or MEF2C was acetylated. Mutant Myb-K456A exhibited reduced acetylation and weak stimulation for lncAY transcription. However, Myb mutation K456/503A prevented Myb from acetylation induced by sulfatide. The mutant Myb K456/503A further was unable to occupy lncAY promoter and enhance lncAY transcription. In conclusion, this study demonstrated lncAY transcription was abnormally upregulated by sulfatide in HCC through Myb/MEF2C to promote HCC progression.

LncRNA AY promotes hepatocellular carcinoma metastasis by stimulating ITGAV transcription.

Rationale: Tumor metastasis is the main cause for cancer-related death. However, the driving molecules of metastasis remain largely unknown. Here, we aim to identify long non-coding RNAs (lncRNAs) critical for human hepatocellular carcinoma (HCC) metastasis. Methods: Microarrays were used to screen a comprehensive set of lncRNAs with differential expression profiles in sulfatide-treated cells. Mass spectrometry, protein arrays, and RNA pull-down experiments were used to identify proteins that interacted with lncRNA. Epigenetic analysis was used to study lncRNA-mediated regulation mechanisms. Results: We identified lncRNA AY927503 (AY) as a metastasis-associated molecule that was highly expressed in human hepatocellular carcinoma (HCC) and correlated with metastatic events and poor prognosis in patients with HCC. AY promoted HCC cell migration, stemness, 5-fluorouracil resistance, and metastasis in mice. However, knockdown of integrin αV (ITGAV) abolished AY-stimulated migration, cell viability in HCC cells or tube formation. AY strongly promoted ITGAV transcription and αVß3 expression by interacting with the ITGAV promoter specifically and stimulating its activity. AY was identified to interact with histone 1FX (H1FX), but deletion of the central domain of AY (AY∆371-522) abolished H1FX binding and ITGAV promoter stimulation. AY significantly enriched H3K4Me3 and acH3K9/14 but reduced H3K27Me3 and H1FX occupancy on the ITGAV promoter, which remodeled chromatin structures for RNA polymerase II recruitment. Knockdown of H1FX abrogated ITGAV transcription stimulated by AY. Conclusions: Our findings suggested that lncRNA AY promoted HCC metastasis via induction of chromatin modification for ITGAV transcription as a pioneer factor and was a potential molecular signature for metastasis or poor prognosis in patients with HCC.

BRD1-Mediated Acetylation Promotes Integrin αV Gene Expression Via Interaction with Sulfatide.

Integrin αV gene expression is often dysregulated in cancers especially in hepatocellular carcinoma (HCC); however, the mechanism of regulation is poorly understood. Here, it is demonstrated that sulfatide activated integrin αV gene transcription, through histone H3K9/14 acetylation at the promoter, and high integrin αV expression are closely associated with poor prognosis. To elucidate the mechanism of regulation of acetylation, sulfatide-bound proteins were screened by mass spectrometry (MS), and bromodomain containing protein 1 (BRD1) was identified as an interacting protein that also colocalized with sulfatide in HCC cells. BRD1 was also formed a complex with Sp1, which was recruited to the integrin αV gene promoter. Sulfatide was also found to induce BRD1, monocytic leukemia zinc finger (MOZ) and histone acetyltransferase binding to ORC1 (HBO1) acetyltransferase multiprotein complex recruitment to the integrin αV promoter, which is responsible for histone H3K9/14 acetylation. Finally, knockdown of BRD1 limited sulfatide-induced H3K9/14 acetylation and occupancy of MOZ or HBO1 on integrin αV gene promoter.Implications: This study demonstrates that sulfatide interaction with BRD1 mediates acetylation and is important for regulation of integrin αV gene expression. Mol Cancer Res; 16(4); 610-22. ©2018 AACR.

Nr3C1-Bhlhb2 Axis Dysregulation Is Involved in the Development of Attention Deficit Hyperactivity.

Attention deficit hyperactivity disorder (ADHD) is a child developmental and behavioral disorder which seriously hinders their education and development. To investigate the key regulators in the prefrontal cortex (PFC), the major affected areas of ADHD, microRNA (miR)-138,138*, 34c*, 296, and 494, were noted for their significant downregulation in ADHD model rats spontaneously hypertensive rats (SHRs) compared to Wistar Kyoto (WKY) rat control. Based on promoter sequence analysis and activity assay, glucocorticoid receptor (Nr3c1) was identified for the inhibition of the promoter activity of miR-138-1, 34c*, 296, and 494 genes and their transcription. In the PFC of ADHD model rats SHR, Nr3c1 expression was abnormally elevated and reversely correlated with the levels of miR-138-1, 34c, 296, and 494 expression. Luciferase report assays indicated that all miR-138, 138*, 34c*, 296, and 494 targeted the 3' untranslated region of transcription factor Bhlhb2 (Bhlhe40) messenger RNA (mRNA) in common and ectopic expression of miR-138,138*, 34c*, 296, and 494 further suppressed the expression of Bhlhb2 gene. Consistently, Bhlhb2 expression was significantly higher in PFC of ADHD model SHR than control. Overexpressed Bhlhb2 in vitro significantly suppressed PC12 cell differentiation, and silence of Bhlhb2 enhanced the growth of neurite axon and dendrite. To observe the roles of Bhlhb2 further in vivo, Bhlhb2 was silenced in the PFC of nine SHR rats. Interestingly, knockdown of Bhlhb2 significantly improved the hyperactivity behaviors in SHRs compared to control. These findings show that Nr3c1-Bhlhb2 axis dysregulation was involved in the development of attention deficit and hyperactivity.

MiR-124 inhibits the migration and invasion of human hepatocellular carcinoma cells by suppressing integrin αV expression.

Tumor metastasis is the major cause of cancer-related death especially in human hepatocellular carcinoma (HCC). Although microRNAs have been implicated in tumor development, the roles of miR-124 in HCC metastasis are still not well understood. We conducted functional analysis in this study to investigate miR-124. We observed that miR-124 significantly retarded the wound healing and migration of HCC SMMC-7721 and BEL-7404 cells. Further analysis indicated miR-124 directly targeting the transcriptional factor Sp1 which is an important transcription factor for the integrin αV subunit gene transcription. Co-transfection of miR-124 with the luciferase reporter containing Sp1 3' untranslated region (UTR) significantly suppressed the luciferase activities. While mutation of the binding site of miR-124 in Sp1 mRNA 3'UTR completely abrogated the suppression of miR-124. Overexpression of miR-124 resulted in robust downregulation of Sp1 and integrin αV expression at either mRNA or protein level. Ectopic expression of miR-124 in HCC dramatically repressed the wound healing and migration in vitro and tumor metastasis in mouse experiments. Our findings demonstrated that miR-124 played as an important role in regulation of integrin αV expression in HCC, and reintroduction of miR-124 might be an alternative therapeutic strategy for controlling integrin αV expression in HCC.

Remarkable expression of vascular endothelial growth factor in bone marrow plasma cells of patients with POEMS syndrome.

Vascular endothelial growth factor (VEGF) is pathognomonically elevated in patients with POEMS (polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy and skin changes) syndrome. However, its source of overproduction is unclear. As clinical improvement is almost always associated with VEGF reduction after anti-plasma cell therapy, its increase at diagnosis has been attributed to the underlying monoclonal gammopathy, although direct evidence is still lacking. In the current study, we systemically measured VEGF levels in POEMS patients, before and after treatment. Bone marrow plasma cells showed remarkable VEGF expression, in both mRNA and protein levels, which decreased gradually in response to therapy. Of note, statistically linear correlations were observed between serum and bone marrow plasma cell VEGF levels (mRNA vs. serum, rho 0.343, p=0.003; protein vs. serum, rho 0.644, p<0.0001), supporting bone marrow plasma cells as the main source of circulating VEGF. Intriguingly, immunophenotyping revealed that bone marrow plasma cells were polyclonal in most patients at diagnosis. A clear monoclonal population, coexistent with polytypic cells, was only detectable in 11 cases (18%), in which comparable intracellular VEGF expression was observed between these two plasma cell populations (p=0.594), while monoclonal cells showed higher intracellular interleukin-6 expression (p=0.006). These patients had more serum monoclonal protein, less post-therapeutic complete remission, and inferior overall (p=0.027) and progression-free survival (p=0.002). Collectively, bone marrow plasma cells, mainly polyclonal population, are the major source of VEGF overproduction in POEMS patients.

Rapidly Progressive Polyneuropathy in a Patient With Monoclonal Gammopathy: A Case Report of POEMS Syndrome and Beyond.

Neuropathy, the dominant clinical feature of POEMS syndrome, is typically distal, symmetric, and slowly progressive with demyelinating changes. After a gradual proximal spread, it usually results in severe muscle weakness and functional disabilities. Cases characterized by acute onset polyneuropathy are rarely described. In the present report, we describe a 32-year-old male diagnosed as POEMS syndrome, but presenting with a rapidly evolving polyneuropathy. Detailed clinical, electrophysiological, and genetic studies revealed a coexisting underdiagnosed inherited axonal neuropathy, namely Charcot-Marie-Tooth disease 2A2. The patient received lenalidomide-based chemotherapy and consolidated by autologous stem cell transplantation for his POEMS syndrome, which improved the neurological disability. In most conditions, only 1 cause is responsible for a patient's polyneuropathy. However, an insidious inherited neuropathy can be overlooked, when an acquired condition is present. The case illustrated here, to the best of our knowledge, is the first one with coexistent axonal type Charcot-Marie-Tooth disease and POEMS syndrome, suggesting that an unrecognized inherited neuropathy may change the disease course of a further acquired neuropathy.

Sulfatide interacts with and activates integrin αVß3 in human hepatocellular carcinoma cells.

Integrin αVß3 is a malignant driver of anchorage-independence and tumor angiogenesis, but its dysregulation in hepatocellular carcinoma (HCC) remains unclear. In this study, we observed that sulfatide significantly promoted integrin αV(ITGAV) expression and wound closure in HCC. We also noted that elevated sulfatide profoundly stimulated integrin αVß3 clustering and signaling. In the cells with integrin αVß3 clustering induced by sulfatide, integrin ß3 subunit was phosphorylated. Simultaneously, focal adhesion kinase (FAK), Src and paxillin were also phosphorylated. Treatment with FAK inhibitor resulted in robust suppression of FAK-Y397 and Src-Y416 phosphorylation stimulated by sulfatide, but not suppression of integrin ß3 phosphorylation. Src inhibitors repressed Src-Y416 and FAK Y861 and Y925 phosphorylation, but not FAK-Y397 and integrin ß3 phosphorylation. After mutation of integrin ß3 (Y773F and Y785F), FAK or Src phosphorylation failed to be stimulated by sulfatide. Moreover, ß3 Y773 and Y785 phosphorylation was suppressed by insulin-like growth factor receptor knockdown even in cells stimulated by sulfatide. In assays of immunoprecipitation and immunostaining with integrin αV or ß3 antibody, labeled sulfatide was found in the complex and co-localized with integrin αVß3. Taken together, this study demonstrated that elevated sulfatide bound to integrin αVß3 and induced clustering and phosphorylation of αVß3 instead of matrix ligand binding, triggering outside-in signaling.