A novel mutation in ext2 caused hereditary multiple exostoses through reducing the synthesis of heparan sulfate.

Hereditary multiple exostoses (HME) is a rare skeletal disorder characterized by the formation of multiple benign cartilage-capped tumors, usually in the metaphyseal region of the long bones. Over 70% of HME cases arise from monoallelic mutations in either of the two genes encoding the heparan sulfate (HS) synthesis enzymes, ext1 and ext2. To identify more HME-associated mutations, genomic DNA from members of five independent consanguineous families with HME was sequenced with whole exome sequencing (WES). A novel heterozygous splice site mutation (c.1173+2T>A) in ext2 was detected in all three affected members of family V. Further study showed that the novel mutation caused exon 7 of ext2 mRNA to be skipped during splicing and caused a frameshift after the codon for Arg360, which results in the appearance of new 43 codons, followed by a termination codon. Although the resulting truncated protein was still localized to the Golgi, similar to the full-length EXT2, its HS synthesis activity decreased by 40%. In this study, a novel splice site mutation in ext2 was identified and suggested to be a pathogenic mutation of HME, which may expand the genetic etiology spectrum of HME and may be helpful for clinical genetic counseling and prenatal diagnosis.

Exosomal-lncRNA DLEU1 Accelerates the Proliferation, Migration, and Invasion of Endometrial Carcinoma Cells by Regulating microRNA-E2F3.

PURPOSE: Long non-coding RNAs (lncRNAs) may act as oncogenes in several cancers, including endometrial carcinoma (EC). The purpose of the current study is to investigate the regulatory mechanism of exosomal-lncRNA deleted in lymphocytic leukemia1 (DLEU1) on EC. METHODS: The expression levels of lncRNA DLEU1, microRNA-381-3p and E2F Transcription Factor 3 (E2F3) in EC tissues or cells were detected using quantitative reverse transcription-polymerase chain reaction (qRT-PCR). We then analysed the proliferation, migration, and invasion of EC cells by performing the MTT assay, wound healing assay, and transwell invasion assay, respectively. Identification of exosomes was detected using Western blot assay. The uptake of exosomes was detected by a confocal microscope. The effects of exosomes on EC cells were investigated by construction of cell co-culture system. The interactions among DLEU1, miR-381-3p and E2F3 were confirmed using the dual-luciferase reporter (DLR) assay. RESULTS: LncRNA DLEU1 expression was highly up-regulated in EC tissues and cells. Knockdown of DLEU1 inhibited the proliferation, migration, and invasion of EC cells. Exosomes could be uptaken by the ambient EC cells. MiR-381-3p was a target of DLEU1 and was negatively modulated by DLEU1. Overexpression of miR-381-3p suppressed the proliferation, migration, and invasion of EC cells. Additionally, E2F3 was the target gene of miR-381-3p and was negatively modulated by miR-381-3p. Upregulation of miR-381-3p and down-regulation of E2F3 reversed the promoting effect of exosomal DLEU1 on EC cells. CONCLUSION: Exosomal DLEU1 accelerates the development of EC by regulating the miR-381-3p/E2F3 axis, thus DLEU1 may act as a possible therapeutic target for treating EC.

Advances in next-generation lipid-polymer hybrid nanocarriers with emphasis on polymer-modified functional liposomes and cell-based-biomimetic nanocarriers for active ingredients and fractions from Chinese medicine delivery.

Recently, functional liposomes modified with versatile polymer and cell-based- biomimetic nanoparticles have emerged as the most advanced lipid-polymer hybrid nanocarriers (LPNs) for drug delivery. This review highlights the advances of these two LPNs in the delivery of active ingredients and fractions from Chinese medicine with promising therapeutic, chemopreventive, or chemosensitive effects. To understand their complete potency, the relationship between the nanoparticle characteristics and their in vitro and in vivo performance characteristics has been discussed. Polymer-modified liposomes and cell-based biomimetic nanoparticles are beneficial for improving absorption, modulating release, targeting and overcoming multidrug resistance, and reducing side effects. The associated challenges, current limitations, and opportunities in this field are also discussed.

Functional oral nanoparticles for delivering silibinin and cryptotanshinone against breast cancer lung metastasis.

BACKGROUND: Breast cancer lung metastasis occurs in more than 60% of all patients with breast cancer, and most of those afflicted by it eventually die of recurrence. The tumor microenvironment plays vital roles in metastasis. Modulating the tumor microenvironment via multiple pathways could efficiently prevent or inhibit lung metastasis. Silibinin and cryptotanshinone are natural plant products that demonstrate anti-metastasis effects and modulate the tumor microenvironment via different pathways. However, they have poor aqueous solubility, membrane permeability, and oral bioavailability. Oral drug administration may help improve the quality of life and compliance of patients with breast cancer, primarily under long-term and/or follow-up therapy. Herein, we developed poly-N-(2-hydroxypropyl) methacrylamide (pHPMA)-coated wheat germ agglutinin-modified lipid-polymer hybrid nanoparticles, co-loaded with silibinin and cryptotanshinone (S/C-pW-LPNs). We assessed their oral bioavailability, and evaluated their anti-metastasis efficacy in a 4T1 breast cancer tumor-bearing nude mouse model. RESULTS: An in vitro mucus diffusion study revealed that pHPMA enhanced W-LPN mucus penetration. After oral administration, pHPMA enhanced nanoparticle distribution in rat jejunum and substantially augmented oral bioavailability. S/C-W-LPNs markedly increased 4T1 cell toxicity and inhibited cell invasion and migration. Compared to LPNs loaded with either silibinin or cryptotanshinone alone, S/C-pW-LPNs dramatically slowed tumor progression in 4T1 tumor-bearing nude mice. S/C-pW-LPNs presented with the most robust anti-metastasis activity on smooth lung surfaces and mitigated lung metastasis foci. They also downregulated tumor microenvironment biomarkers such as CD31, TGF-ß1, and MMP-9 that promote metastasis. CONCLUSIONS: Silibinin- and cryptotanshinone-co-loaded pW-LPNs efficiently penetrate intestinal barriers, thereby enhancing the oral bioavailability of the drug loads. These nanoparticles exhibit favorable anti-metastasis effects in breast cancer-bearing nude mice. Hence, S/C-pW-LPNs are promising oral drug nanocarriers that inhibit breast cancer lung metastasis.

MicroRNA-155 Suppresses the Translation of p38 and Impairs the Functioning of Dendritic Cells in Endometrial Cancer Mice.

BACKGROUND: Dendritic cells (DCs) are reported to play an important role in activating the anti-tumor immune responses. p38 MAPK14 signaling plays an important role in controlling their activity. Here, we identified that miR-155 suppressed the translation of p38 and impaired the functioning of dendritic cells in endometrial cancer. METHODS: HEC1A endometrial cancer cell lines were used for the study which was transfected in the C57BL/6 mice. Murine bone marrow-derived dendritic cells (BMDCs) were isolated from the mice. Target prediction was done by TargetScan which was confirmed by RT-PCR analysis. The protein expression was carried by Western blot analysis. Levels of IL-12 were evaluated by ELISA. Mice injected with HEC1A cells were subjected to tumor challenge study. RESULTS: On screening the binding sites of p38 MAPK14 gene, miR-155 was found to bind the 3'UTR directly and blocked its translation. The levels of miR-155 were upregulated in dendritic cells and RAW264.7 cells, miR-155 showed inhibitory effect on expression levels of p38. In dendritic cells, miR-155 was found to regulate the expression of IL-12, also miR-155 inhibitor stimulated the differentiation of Th1 cells in mice induced with endometrial cancer. In dendritic cells, miR-155 inhibited the expression of p38 gene and decreased their ability to interfere in tumor growth. CONCLUSION: The study concludes suppressive role of miR-155 in the process of dendritic cells mediated anti-tumor immunity, also inhibiting miR-155 provides a novel strategy for countering endometrial cancer.

Functional lipid polymeric nanoparticles for oral drug delivery: Rapid mucus penetration and improved cell entry and cellular transport.

To improve Biopharmaceutics Classification System class IV drug bioavailability, mucus and underlying intestinal epithelial barriers must be overcome. Hydrophilic nanoparticle coatings may hinder cellular uptake and transport. We integrated hydrophilic, detachable poly(N-(2-hydroxypropyl) methacrylamide) with vitamin B12-modified chitosan into lipid polymeric nanoparticles (H/VC-LPNs) to enhance mucus penetration, intracellular uptake, and transepithelial absorption. Multiple particle tracking revealed accelerated mucus diffusion into porcine mucus in vitro. The nanoparticles increased uptake and intracellular distribution in Caco-2 cells, which may involve intrinsic factor receptor-mediated endocytosis and intercellular tight junctions. Integration of improved mucus penetration and intracellular absorption was confirmed by in vitro internalization kinetics in HT29-MTX/Caco-2 co-cultures and in vivo distribution, transport, and mouse Peyer's patch absorption. H/VC-LPNs substantially increased curcumin bioavailability in rats. A nanocarrier with a dissociable shell, receptor-mediated intracellular penetration, and paracellular transport may be promising for oral curcumin delivery. This study identified the key factors involved in oral bioavailability enhancement.

Hybrid curcumin-phospholipid complex-near-infrared dye oral drug delivery system to inhibit lung metastasis of breast cancer.

Background: Oral route of administration is preferred for treating breast cancer, especially when continued disease management with good tolerability is required; however, orally administered chemotherapeutics combined with near-infrared (NIR) dyes are hindered by the low bioavailability, insufficient for the desired therapeutic efficacy. In this study, we developed a hybrid self-microemulsifying drug delivery system for co-loading curcumin-phospholipid complex and NIR dye IR780 (CUR/IR780@SMEDDS), to achieve combined phototherapeutic and chemotherapeutic effects against lung metastasis of breast cancer. Methods: CUR/IR780@SMEDDS were characterized. The efficacy against breast cancer metastasis was evaluated by photothermal and photodynamic assessment, cytotoxicity, invasion, and migration in metastatic 4T1 breast cancer cells in vitro, and in vivo oral bioavailability study in rats and pharmacodynamics studies in tumor-bearing nude mice. Results: CUR/IR780@SMEDDS improved oral bioavailability of curcumin and IR780 in rats compared with curcumin and IR780 suspensions. CUR/IR780@SMEDDS exhibited remarkable photothermal and photodynamic effects in vitro. In metastatic 4T1 breast cancer cells, CUR/IR780@SMEDDS combined with localized NIR laser irradiation induced significant cytotoxicity and inhibited invasion and migration of 4T1 cells, an outcome attributable to cumulative effects of IR780-induced hyperthermia and pharmacological effects of curcumin. In orthotopic 4T1 tumor-bearing nude mice, combination of oral administration of CUR/IR780@SMEDDS with local NIR laser irradiation inhibited tumor progression and suppressed lung metastasis.

A Krüppel-Like Factor 1 Gene Mutation Ameliorates the Severity of ß-Thalassemia: A Case Report.

Patients with the ß0/ß0 type of ß-thalassemia (ß-thal) usually present as ß-thal major (ß-TM), and are transfusion-dependent. However, the clinical and hematological features of ß-thal can be modulated by different modifiers, resulting in a wide range of clinical severity even in patients with the same genotypes. We report a Chinese family with twin brothers, both of whom had the same genotype of ß0/ß0. One twin was diagnosed as ß-TM at 4 months of age and had regularly been transfused; conversely the other twin with a KLF1 (Krüppel-like factor 1) gene mutation, behaved as ß-thal intermedia (ß-TI), and had never been transfused. Our findings indicate that KLF1 mutations have a role in modulating the phenotypic severity of ß-thal. The exact investigation of KLF1 modifiers is necessary in areas where globin gene disorders are most prevalent. This will be helpful in genetic counseling and optimizing the guidelines for prenatal diagnosis (PND) programs.

Pre Gestational Thalassemia Screening in Mainland China: The First Two Years of a Preventive Program.

In this study, we report the experience of a pre gestational thalassemia screening program at a single center in Southern China. Free thalassemia screening, genetic counseling and prenatal diagnosis (PND) for couples planning pregnancy were implemented over a 2-year period. Among a total of 83,062 screened individuals (41,531 couples), the allele frequencies of ß-thalassemia (ß-thal), - -SEA and - -THAI deletions were 3.79, 5.75 and 0.028%, respectively. Out of the 41,531 couples, 11,039 couples had at least one partner who had a positive screening test; of these, 455 at-risk couples (1.07%) were identified, including 68 (0.16%) for ß-thal, 162 (0.39%) for Hb Bart's (γ4) hydrops fetalis, 190 (0.46%) for deletional Hb H (ß4) disease and 25 (0.06%) for nondeletional Hb H disease. Of the 455 at-risk couples, 90 were already pregnant and 66 underwent PND at 10-13 weeks' gestation, resulting in 15 affected fetuses. The remaining 355 at-risk couples were still preparing for pregnancy, and they were on the list for follow-up. There is considerable scope for facilitating timely PND through improved organization and screening strategy. The pre pregnancy screening is a feasible and effective approach to thalassemia prevention.

A Novel Frameshift Mutation at Codons 138/139 (HBB: c.417_418insT) on the ß-Globin Gene Leads to ß-Thalassemia.

We describe a new ß-thalassemic mutation in a Chinese subject. This allele develops by insertion of one nucleotide (+T) between codons 138 and 139 in the third exon of the ß-globin gene. The mutation causes a frameshift that leads to a termination codon at codon 139. In the heterozygote, this allele has the phenotype of classical ß-thalassemia (ß-thal) minor.

Hb Hornchurch [ß43(CD2)Glu→Lys; HBB: c.130G>A] Compromises the Molecular Diagnosis of ß-Thalassemia in a Chinese Family.

The combination of ß-thalassemia (ß-thal) and a hemoglobin (Hb) variant is not uncommon in regions with a high prevalence of thalassemia. Although most of the ß-globin chain variants will not aggravate the ß-thal, some can compromise the accurate molecular diagnosis. In this study, we present a rare case of coinheritance of ß-thal and Hb Hornchurch [ß43(CD2)Glu→Lys; HBB: c.130G>A], that compromises the molecular diagnosis of homozygous ß-thal.

Hydrops Fetalis Associated with Compound Heterozygosity for Hb Zurich-Albisrieden (HBA2: C.178G > C) and the Southeast Asian (- -SEA/) Deletion.

Hb Zurich-Albisrieden [HBA2: c.178G > C; α59(E8)Gly→Arg (α2)] is a rare nondeletional α-thalassemia (α-thal) that results from a nucleotide substitution at codon 59 of the α2-globin gene. In this report, we present a fetus with cardiomegaly, enlarged placenta and increased middle cerebral artery-peak systolic velocity (MCA-PSV) at 25 weeks' gestation. Fetal blood sampling revealed the severe anemia [hemoglobin (Hb) level being 5.5 g/dL] and Hb H (ß4) disease-like hematological findings with Hb Bart's (γ4) level of 30.7%. Molecular analysis of the family found that the father was an Hb Zurich-Albisrieden carrier, the mother heterozygous for the - -SEA α0-thal deletion, and the fetus was a compound heterozygote for Hb Zurich-Albisrieden and the - -SEA α0-thal deletion. Therefore, this was a rare case of Hb Bart's hydrops fetalis associated with Hb Zurich Albisrieden.

Diagnostic Dilemma of Hb Perth [ß32(B14)Leu→Pro; HBB: c.98T > C] in Mainland China.

Unstable hemoglobin (Hb) variants represent a rare etiology of congenital hemolytic anemia. Correct diagnosis can be a challenge due to the relative rarity or lack of awareness of this disorder. We report an 18-month-old girl, who presented with a long-standing hemolytic anemia. Her diagnosis of unstable Hb Perth [ß32(B14)Leu→Pro, HBB: c.98T > C] had not been made until gene sequencing of the ß-globin gene was performed.

A New δ-Globin Gene Variant: Hb A2-Fengshun [δ121(GH4)Glu→Lys (HBD: c.364G > A)].

An elevated Hb A2 (α2δ2 level) is a diagnostic marker for heterozygous ß-thalassemia (ß-thal). Mutations in the δ-globin gene can cause decreased expression of Hb A2, compromising screening for heterozygous ß-thal. In this report, we describe a novel missense mutation of the δ-globin [Hb A2-Fengshun or δ121(GH4)Glu→Lys, HBD: c.364G > A] in a Chinese individual who had coinherited a heterozygous ß-thal with a normal Hb A2 level.

Consequences of Delayed Prenatal Diagnosis of ß-Thalassemia in Mainland China.

ß-Thalassemia (ß-thal) is one of the most common inherited single gene disorders in the world. The aim of this study was to describe the gestational age at prenatal diagnosis (PND) for ß-thal in at-risk women in mainland China. All pregnant women at-risk for ß-thal and undergoing PND at a Mainland Chinese tertiary obstetric center between January 2005 and December 2014 were included. Information required for the survey was obtained from prenatal records and delivery charts. In total, 1307 women underwent PND for ß-thal. The mean gestational age for the procedure was 18.5 weeks. There were 384 (29.0%) women with fetal diagnosis in early trimester (<14 weeks), 715 (55.0%) in early second trimester (14-24 weeks), and 208 (16.0%) in late second trimester or beyond (>24 weeks). Although the proportion of patients undergoing early PND increased along with the time span, the mean n gestational age was not decreased significantly during the study period. The delay in PND deprived couples of the opportunity to make informed decisions early in pregnancy.

Drought monitoring based on TIGGE and distributed hydrological model in Huaihe River Basin, China.

Drought assessment is important for developing measures to reduce agricultural vulnerability and thereby secure the livelihoods of those who depend on agriculture. This study uses four global ensemble weather prediction systems: the China Meteorological Administration (CMA), the European Centre for Medium-Range Weather Forecasts (ECMWF), the UK Met Office (UKMO), and the US National Centres for Environmental Prediction (NCEP) in the THORPEX (The Observing System Research and Predictability Experiment) Interactive Grand Global Ensemble (TIGGE) archive from 2006 to 2010. Based on results from the XXT (the first X denotes Xinanjiang, the second X denotes hybrid, and the T denotes TOPMODEL) distributed hydrological model, as well as soil moisture observations and digital elevation model (DEM) data, synthesized drought grades were established in the Huaihe River Basin of China. To filter out the impact of short-term fluctuations on observed soil moisture, a 30-day moving average was calculated. Use of the moving average significantly improves the correlation between observed soil moisture and simulated soil water deficit depth. Finally, a linear regression model describing the relationship between observed soil moisture and simulated soil water deficit depth was constructed. The deterministic regression coefficient was 0.5872, the correlation coefficient was 0.77, and the regression coefficient was -154.23. The trends in drought grades calculated using soil moisture and soil water deficit depth were found to be the same, and the grades agreed to within one level. Our findings highlight the importance of synthesizing drought grading when assessing drought using different soil moisture indicators in order to obtain a more comprehensive forecast of drought conditions.

Evidence of Selection for the α-Globin Gene Deletions and Triplications in a Southern Chinese Population.

α(+)-Thalassemia (α(+)-thal) is common in Southern China. The high frequency could be due to over dominant selection through malaria. Two molecular mechanisms that produce α(+)-thal have been defined; one results in the -α(3.7) (rightward) deletion and reciprocal ααα(anti 3.7) triplication, and the other one results in the -α(4.2) (leftward) deletion and reciprocal ααα(anti 4.2) triplication. Considering that each de novo event produced a chromosome with an α gene deletion and a chromosome with an α triplication, if there is no favorable allele, one would expected to find the same allelic frequencies. We found a favorable selection for the -α(3.7) deletion in the Chinese population, and we also found that the α triplication is not as rare as was first thought, especially for the ααα(anti 3.7) triplication.