Simultaneously degradation of various phthalate esters by Rhodococcus sp. AH-ZY2: Strain, omics and enzymatic study.

Phthalate esters (PAEs) are widely used as plasticizers and cause serious complex pollution problem in environment. Thus, strains with efficient ability to simultaneously degrade various PAEs are required. In this study, a newly isolated strain Rhodococcus sp. AH-ZY2 can degrade 500 mg/L Di-n-octyl phthalate completely within 16 h and other 500 mg/L PAEs almost completely within 48 h at 37 °C, 180 rpm, and 2 % (v/v) inoculum size of cultures with a OD600 of 0.8. OD600 = 0.8, 2 % (v/v). Twenty genes in its genome were annotated as potential esterase and four of them (3963, 4547, 5294 and 5359) were heterogeneously expressed and characterized. Esterase 3963 and 4547 is a type I PAEs esterase that hydrolyzes PAEs to phthalate monoesters. Esterase 5294 is a type II PAEs esterase that hydrolyzes phthalate monoesters to phthalate acid (PA). Esterase 5359 is a type III PAEs esterase that simultaneously degrades various PAEs to PA. Molecular docking results of 5359 suggested that the size and indiscriminate binding feature of spacious substrate binding pocket may contribute to its substrate versatility. AH-ZY2 is a potential strain for efficient remediation of PAEs complex pollution in environment. It is first to report an esterase that can efficiently degrade mixed various PAEs.

The bacterial patterns suggesting the dynamic features of tick-associated microorganisms in hard ticks.

BACKGROUND: Ticks are blood-feeding significant arthropods that can harbour various microorganisms, including pathogens that pose health risks to humans and animals. Tick-symbiont microorganisms are believed to influence tick development, but the intricate interactions between these microbes and the relationships between different tick-borne microorganisms remain largely unexplored. RESULTS: Based on 111 tick pool samples presenting questing and engorged statuses including 752 questing tick and 1083 engorged tick from cattle and goats, which were collected in two types of geographic landscape (semi-desert and alpine meadow). We observed significant variations in the composition of tick-borne microorganisms across different environments and blood-engorgement statuses, with a pronounced divergence in symbionts compared to environmental bacteria. Metabolic predictions revealed over 90 differential pathways for tick-borne microorganisms in distinct environments and more than 80 metabolic variations in response to varying blood engorgement statuses. Interestingly, nine pathways were identified, particularly related to chorismate synthesis and carbohydrate metabolism. Moreover, microbial network relationships within tick-borne microorganism groups were highly distinct across different environments and blood-engorgement statuses. The microbial network relationships of symbionts involve some pathogenic and environmental microorganisms. Regression modelling highlighted positive correlations between the Coxiella symbiont and related pathogens, while some environmental bacteria showed strong negative correlations with Coxiella abundance. We also identified commensal bacteria/pathogens in bacterial cooccurrence patterns. Furthermore, we tested pathogenic microorganisms of each tick sample analysis revealed that 86.36% (1601/1855) of the tick samples carried one or more pathogenic microorganisms, The total carrier rate of bacterial pathogens was 43.77% ((812/1855). Most blood samples carried at least one pathogenic microorganism. The pathogens carried by the ticks have both genus and species diversity, and Rickettsia species are the most abundant pathogens among all pathogens. CONCLUSION: Our findings underscore that the bacterial pattern of ticks is dynamic and unstable, which is influenced by the environment factors and tick developmental characteristics.

Macrocycle-Based Charge Transfer Cocrystals with Dynamically Reversible Chiral Self-Sorting Display Chain Length-Selective Vapochromism to Alkyl Ketones.

Chirality-driven self-sorting plays an essential role in controlling the biofunction of biosystems, such as the chiral double-helix structure of DNA from self-recognition by hydrogen bonding. However, achieving precise control over the chiral self-sorted structures and their functional properties for the bioinspired supramolecular systems still remains a challenge, not to mention realizing dynamically reversible regulation. Herein, we report an unprecedented saucer[4]arene-based charge transfer (CT) cocrystal system with dynamically reversible chiral self-sorting synergistically induced by chiral triangular macrocycle and organic vapors. It displays efficient chain length-selective vapochromism toward alkyl ketones due to precise modulation of optical properties by vapor-induced diverse structural transformations. Experimental and theoretical studies reveal that the unique vapochromic behavior is mainly attributed to the formation of homo- or heterochiral self-sorted assemblies with different alkyl ketone guests, which differ dramatically in solid-state superstructures and CT interactions, thus influencing their optical properties. This work highlights the essential role of chiral self-sorting in controlling the functional properties of synthetic supramolecular systems, and the rarely seen controllable chiral self-sorting at the solid-vapor interface deepens the understanding of efficient vapochromic sensors.

An improved medium for in vitro studies of female reproduction and oviposition in Schistosoma japonicum.

BACKGROUND: Schistosomiasis is a disease primarily caused by eggs laid by pathogens called schistosomes. Among the schistosome species infecting humans, Schistosoma japonicum possesses the largest fecundity; each adult female produces an average of 3500 eggs per day. The lack of proper culture conditions supporting continuous oviposition in vitro has precluded detailed investigation of mechanisms regulating sexual maturation and egg production in Schistosoma japonicum. METHODS: We optimized in vitro culture conditions by replacing reagents that are part of the classical ABC169 medium. Fast Blue BB staining and 4',6-diamidino-2-phenylindole (DAPI) labeling were applied to observe the sexual development status of the females. In vitro RNA interference (RNAi) technology was used to validate the capability of the modified medium. The detection of male ß-alanyl-tryptamine (BATT) was conducted using liquid chromatography-mass spectrometry (LC-MS). RESULTS: Both m-AB169 (1640) and AB169 (1640) media are capable of facilitating the sexual development of paired virgin female S. japonicum, as well as sustaining the mature reproductive organs and egg production of adult S. japonicum for at least 22 days in vitro. M-AB169 (1640) provided a more stable condition for supporting the sexual maturity of female S. japonicum, as evidenced by the consistent initiation of egg production compared with AB169 (1640). Through a comparative analysis of S. japonicum and S. mansoni in diverse media, we demonstrated that these closely related species display distinct demands for their sexual development and egg production, suggesting a potential influence of nutritional factors on the observed variations in host ranges among different schistosome species. Importantly, we successfully identified the presence of the pheromone ß-alanyl-tryptamine (BATT) in S. japonicum, previously identified in S. mansoni, highlighting its conserved role in schistosome reproductive development. Through the employment of double-stranded RNA (dsRNA) treatment to silence two genes that are involved in either the male (gli1, glioma-associated oncogene homolog 1) or female (vf1, vitellogenic factor 1) side in male-induced female reproductive development of S. mansoni, we confirmed that the combination of m-AB169 (1640) and RNAi technology has the capacity to facilitate in vitro studies of S. japonicum's reproductive and oviposition processes. CONCLUSIONS: We developed a novel medium, m-AB169 (1640), that not only maintains the mature reproductive organs and continuous oviposition of adult female Schistosoma japonicum for up to 22 days but also supports the reproductive development and subsequent egg-laying of virgin females after pairing with male worms. This study provides a valuable in vitro platform for functional studies of the mechanisms underlying the fascinating biology of the female sexual development and egg production of S. japonicum, which may accelerate the development of new strategies targeting schistosome egg production.

Assimilation of phthalate esters in bacteria.

The massive usage of phthalate esters (PAEs) has caused serious pollution. Bacterial degradation is a potential strategy to remove PAE contamination. So far, an increasing number of PAE-degrading strains have been isolated, and the catabolism of PAEs has been extensively studied and reviewed. However, the investigation into the bacterial PAE uptake process has received limited attention and remains preliminary. PAEs can interact spontaneously with compounds like peptidoglycan, lipopolysaccharides, and lipids on the bacterial cell envelope to migrate inside. However, this process compromises the structural integrity of the cells and causes disruptions. Thus, membrane protein-facilitated transport seems to be the main assimilation strategy in bacteria. So far, only an ATP-binding-cassette transporter PatDABC was proven to transport PAEs across the cytomembrane in a Gram-positive bacterium Rhodococcus jostii RHA1. Other cytomembrane proteins like major facilitator superfamily (MFS) proteins and outer membrane proteins in cell walls like FadL family channels, TonB-dependent transporters, and OmpW family proteins were only reported to facilitate the transport of PAEs analogs such as monoaromatic and polyaromatic hydrocarbons. The functions of these proteins in the intracellular transport of PAEs in bacteria await characterization and it is a promising avenue for future research on enhancing bacterial degradation of PAEs. KEY POINTS: • Membrane proteins on the bacterial cell envelope may be PAE transporters. • Most potential transporters need experimental validation.

Strategies for Constructing Macrocyclic Arene-Based Color-Tunable Supramolecular Luminescent Materials.

Over the past decades, supramolecular luminescent materials (SLMs) have attracted considerable attention due to their dynamic noncovalent interactions, versatile functions, and intriguing applications in many research fields. From construction to application, great efforts and progress have been made in color-tunable SLMs in recent years. In order to realize multicolor luminescence, various design strategies have been proposed. Macrocyclic chemistry, one of the brightest jewels in the field of supramolecular chemistry, has played a crucial role in the construction of stimuli-responsive and emission-tunable SLMs. Moreover, the flexible and tunable conformation and multiple noncovalent complexation sites of the macrocyclic arenes (MAs) afford a new opportunity to create such dynamic smart luminescent materials. Inspired by our reported work on the color-tunable supramolecular crystalline assemblies modulated by the conformation of naphth[4]arene, this Concept provides a summary of the latest developments in the construction of color-tunable MA-based SLMs, accompanied by the various construction strategies. The aim is to provide researchers with a new perspective to construct color-tunable SLMs with fascinating functions.

Naphth[4]arene: Synthesis, Conformations, and Application in Color-Tunable Supramolecular Crystalline Assemblies.

Although the chemistry of macrocyclic arenes has seen rapid development in recent years, the synthesis of new macrocyclic arenes from aromatic rings with no directing groups remains a challenge. In this work, a new macrocyclic arene, naphth[4]arene (NA[4]A), composed of four naphthalene rings bridged by methylene groups, was synthesized using macrocycle-to-macrocycle conversion. NA[4]A shows 1,3-alternate and 1,2-alternate conformations in the solid state, which can be selectively obtained. By supramolecular co-assembly of NA[4]A and 1,2,4,5-tetracyanobenzene (TCNB) in different concentrations and temperatures, two conformation-dependent crystalline luminescent co-assemblies 1,2-NTC and 1,3-NTC can be selectively prepared. Interestingly, the two charge-transfer crystalline assemblies containing NA[4]A with different conformations show bright yellow and green fluorescence, and also display high photoluminescence quantum yields (PLQYs) of 45 % and 43 %. Furthermore, they exhibit color-tunable two-photon excited upconversion emission.

MicroRNA-1 targets ribosomal protein genes to regulate the growth, development and reproduction of Schistosoma japonicum.

Eggs laid by mature female schistosomes are primarily responsible for the pathogenesis of schistosomiasis and critical for transmission. Consequently, elucidating the mechanism of sexual maturation as well as egg production may lead to new strategies for the control of schistosomiasis. MicroRNAs (miRNAs) are involved in multiple biological processes including reproduction in many organisms, yet their roles have not been well characterized in schistosomes. Here, we investigated microRNA-1 (miR-1), which was downregulated gradually in both male and female Schistosoma japonicum after they reached sexually maturity. The expression of miR-1, as shown with quantitative reverse transcription PCR (qRT-PCR), was lower in the reproductive organs of adult females compared with the somatic tissues. Overexpression of miR-1 in adult worms destroyed the morphological architecture of reproductive organs and reduced the subsequent oviposition, which may be due to the activation of apoptosis pathways. Through in silico analysis, 34 potential target genes of miR-1 were identified, including five ribosomal protein genes, called rp-s13, rp-l7ae, rp-l14, rp-l11 and rp-s24e. In vitro dual-luciferase reporter gene assays and miRNA overexpression experiments further validated that these ribosomal protein genes were directly regulated by miR-1. In contrast to the gene expression of miR-1, qRT-PCR and in situ hybridization experiments demonstrated these ribosomal protein genes were enriched in the sexual organs of adult females. Using RNA interference to silence the ribosomal protein genes in different developmental stages in a mouse model system, we demonstrated that these miR-1 target genes not only participated in the reproductive development of S. japonicum, but also were required for the growth and survival of the parasite in the early developmental stages. Taken together, our data suggested that miR-1 may affect the growth, reproduction and oviposition of S. japonicum by targeting the ribosomal protein genes, which provides insights for exploration of new anti-schistosome strategies.

A metabotropic glutamate receptor affects the growth and development of Schistosoma japonicum.

Schistosomiasis is a zoonotic parasitic disease caused by schistosome infection that severely threatens human health. Therapy relies mainly on single drug treatment with praziquantel. Therefore, there is an urgent need to develop alternative medicines. The glutamate neurotransmitter in helminths is involved in many physiological functions by interacting with various cell-surface receptors. However, the roles and detailed regulatory mechanisms of the metabotropic glutamate receptor (mGluR) in the growth and development of Schistosoma japonicum remain poorly understood. In this study, we identified two putative mGluRs in S. japonicum and named them SjGRM7 (Sjc_001309, similar to GRM7) and SjGRM (Sjc_001163, similar to mGluR). Further validation using a calcium mobilization assay showed that SjGRM7 and SjGRM are glutamate-specific. The results of in situ hybridization showed that SjGRM is mainly located in the nerves of both males and gonads of females, and SjGRM7 is principally found in the nerves and gonads of males and females. In a RNA interference experiment, the results showed that SjGRM7 knockdown by double-stranded RNA (dsRNA) in S. japonicum caused edema, chassis detachment, and separation of paired worms in vitro. Furthermore, dsRNA interference of SjGRM7 could significantly affect the development and egg production of male and female worms in vivo and alleviate the host liver granulomas and fibrosis. Finally, we examined the molecular mechanisms underlying the regulatory function of mGluR using RNA sequencing. The data suggest that SjGRM7 propagates its signals through the G protein-coupled receptor signaling pathway to promote nervous system development in S. japonicum. In conclusion, SjGRM7 is a potential target for anti-schistosomiasis. This study enables future research on the mechanisms of action of Schistosomiasis japonica drugs.

Polymorphisms of Cytochromes P450 and Glutathione S-Transferases Synergistically Modulate Risk for Parkinson's Disease.

Background: Environmental substances such as pesticides are well-known in link with Parkinson's disease (PD) risk. Enzymes including cytochromes P450 (CYPs), esterases and glutathione S-transferases (GSTs) are responsible for the xenobiotic metabolism and may functionally compensate each other for subtypes in the same class. We hypothesize that the genetic effects of each class modulate PD risk stronger in a synergistic way than individually. Methods: We selected 14 polymorphic loci out of 13 genes which encode enzymes in the classes of CYP, esterase, and GST, and recruited a cohort of 1,026 PD and control subjects from eastern China. The genotypes were identified using improved multiplex ligation detection reaction and analyzed using multiple models. Results: A total of 13 polymorphisms remained after Hardy-Weinberg equilibrium analysis. None of the polymorphisms were independently associated with PD risk after Bonferroni correction either by logistic regression or genetic models. In contrast, interaction analyses detected increased resistance to PD risk in individuals carrying the rs12441817/CC (CYP1A1) and rs2070676/GG + GC (CYP2E1) genotypes (P = 0.002, OR = 0.393, 95% CI = 0.216-0.715), or carrying the GSTM1-present, GSTT1-null, rs156697/AG + GG (GSTO2) and rs1695/AA (GSTP1) genotypes (P = 0.003, OR = 0.348, 95% CI = 0.171-0.706). The synergistic effect of GSTs on PD was primarily present in females (P = 0.003). No synergistic effect was observed within genotypes of esterases. Conclusion: We demonstrate a presence of synergistic but not individual impact on PD susceptibility in polymorphisms of CYPs and GSTs. The results indicate that the genetic interplay leads the way to PD development for xenobiotic metabolizing enzymes.

Recent advances on triptycene derivatives in supramolecular and materials chemistry.

Triptycene derivatives, a type of specific aromatic compound, have been attracting much attention in many research areas. Over the past several years, triptycene and its derivatives have been described to be useful and efficient building blocks for the design and synthesis of novel supramolecular acceptors, porous materials and luminescent materials with specific structures and properties. In this review, recent researches on triptycene derivatives in supramolecular and materials chemistry are summarized. Especially, the construction of a new type of macrocyclic arenes and organic cages with triptycene and its derivatives as building blocks are focused on, and their applications in molecular recognition, self-assembly and gas selective sorption are highlighted. Moreover, the applications of triptycene and its derivatives in porous organic materials and thermally activated delayed fluorescence (TADF) materials are also discussed.

Enhanced penetrative siRNA delivery by a nanodiamond drug delivery platform against hepatocellular carcinoma 3D models.

Small interfering RNA (siRNA) can cause specific gene silencing and is considered promising for treating a variety of cancers, including hepatocellular carcinoma (HCC). However, siRNA has many undesirable physicochemical properties that limit its application. Additionally, conventional methods for delivering siRNA are limited in their ability to penetrate solid tumors. In this study, nanodiamonds (NDs) were evaluated as a nanoparticle drug delivery platform for improved siRNA delivery into tumor cells. Our results demonstrated that ND-siRNA complexes could effectively be formed through electrostatic interactions. The ND-siRNA complexes allowed for efficient cellular uptake and endosomal escape that protects siRNA from degradation. Moreover, ND delivery of siRNA was more effective at penetrating tumor spheroids compared to liposomal formulations. This enhanced penetration capacity makes NDs ideal vehicles to deliver siRNA against solid tumor masses as efficient gene knockdown and decreased tumor cell proliferation were observed in tumor spheroids. Evaluation of ND-siRNA complexes within the context of a 3D cancer disease model demonstrates the potential of NDs as a promising gene delivery platform against solid tumors, such as HCC.

Novel Strategy to Combat Antibiotic Resistance: A Sight into the Combination of CRISPR/Cas9 and Nanoparticles.

Antibiotic resistance is a significant crisis that threatens human health and safety worldwide. There is an urgent need for new strategies to control multidrug-resistant (MDR) bacterial infections. The latest breakthrough in gene-editing tools based on CRISPR/Cas9 has potential application in combating MDR bacterial infections because of their high targeting ability to specifically disrupt the drug resistance genes that microbes use for infection or to kill the pathogen directly. Despite the potential that CRISPR/Cas9 showed, its further utilization has been hampered by undesirable delivery efficiency in vivo. Nanotechnology offers an alternative way to overcome the shortcomings of traditional delivery methods of therapeutic agents. Advances in nanotechnology can improve the efficacy and safety of CRISPR/Cas9 components by using customized nanoparticle delivery systems. The combination of CRISPR/Cas9 and nanotechnology has the potential to open new avenues in the therapy of MDR bacterial infections. This review describes the recent advances related to CRISPR/Cas9 and nanoparticles for antimicrobial therapy and gene delivery, including the improvement in the packaging and localizing efficiency of the CRISPR/Cas9 components in the NP (nanoparticle)/CRISPR system. We pay particular attention to the strengths and limitations of the nanotechnology-based CRISPR/Cas9 delivery system to fight nosocomial pathogens.We highlight the need for more scientific research to explore the combinatorial efficacy of various nanoparticles and CRISPR technology to control and prevent antimicrobial resistance.

Mechanism by which the combination of SjCL3 and SjGAPDH protects against Schistosoma japonicum infection.

A vaccine is an important method to control schistosomiasis. Molecules related to lung-stage schistosomulum are considered potential vaccine candidates. We previously showed that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and cathepsin L3 (CL3) displayed differential expression in the lung-stage schistosomula of Schistosoma japonicum cocultured with host cells. In the present study, we prepared the two proteins and detected the protective effects of SjGAPDH by immunizing mice with this protein alone and in combination with SjCL3 with or without Freund's adjuvant. Then, we investigated the possible mechanisms underlying S. japonicum infection. The results showed that vaccination of adjuvanted SjGAPDH decreased the worm burden (37.8%) and egg load (38.1%), and the combination of adjuvanted SjGAPDH and SjCL3 further decreased the worm burden (65.6%) and egg load (70.9%) during Schistosoma japonicum infection. However, the immunization of a combination of adjuvant-free SjGAPDH and SjCL3 displayed a lower protective effect (< 15%) than those of the adjuvanted SjCL3, the adjuvanted SjGAPDH, and a combination of adjuvanted SjGAPDH and SjCL3. Flow cytometric results showed that the frequency of regulatory T cells (Tregs) was lower (P < 0.05) in the group with adjuvanted SjGAPDH and SjCL3 (2.61%) than the remaining groups. The enzyme-linked immunosorbent assay (ELISA) results indicated that except for the uninfected and infected control groups, the remaining groups displayed a Th1-type shift in immune responses. These results showed the immunization of SjGAPDH resulted in partial protection (approximately 38%); inoculation with a combination of SjCL3 and SjGAPDH in Freund's adjuvant resulted in a high immunoprotective effect (> 65%) against Schistosoma japonicum infection in mice, which was possibly caused by the reduced percentage of Tregs and a Th1-type shift in immune responses; and SjCL3 has no adjuvant-like effect, dissimilar to SmCL3.

The genetic variation of different developmental stages of Schistosoma japonicum: do the distribution in snails and pairing preference benefit the transmission?

BACKGROUND: Schistosoma japonicum is a waterborne parasite that causes schistosomiasis in humans and in more than 40 animal species. Schistosoma japonicum shows distinct genetic differentiation among geographical populations and multiple hosts, but the genetic diversity of different developmental stages of S. japonicum from is less studied. Such studies could elucidate ecological mechanisms in disease transmission by analysing feedbacks in individual physiology and population state. METHODS: After infection using cercariae from a pool of snails shedding together (Method I) and infection using mixed equal numbers of cercariae from individually shed snails (Method II), different developmental stages of S. japonicum were genotyped with microsatellite loci, including 346 cercariae, 701 adult worms and 393 miracidia. Genetic diversity and molecular variation were calculated at different population levels. Kinships (I') among cercariae at intra-snail and inter-snail levels were evaluated. Genetic distance (Dsw) was compared between paired and unpaired worms, and partner changing was investigated through paternity identification for miracidia. RESULTS: The cercaria clones in individual snails varied from 1 to 8 and the kinship of cercariae within individual snails was significant higher (P < 0.001) than that among different snails after deleting near-identical multi-locus genotypes (niMLGs). The allelic diversity of worms in Method I was lower (P < 0.001) than that in Method II, and allele frequency among mice in Method I was also less consistent. The parents of some miracidia were worms that were not paired when collected. The Dsw between each female of paired and unpaired males was much larger (P < 0.001) than that between the female and male in each pair. CONCLUSIONS: Most of the infected snails contained multiple miracidia clones. The aggregation of genetically similar S. japonicum miracidia in individual snails and the unbalanced distribution of miracidia among snails suggests a non-uniform genetic distribution of cercariae among snails in the field. This further influenced the genetic structure of adult worms from infections with different cercariae sampling methods. Schistosoma japonicum in mice can change paired partner, preferring to mate with genetically similar worms. These characteristics provide implications for understanding the balance in genetic diversity of S. japonicum related to the transmission of schistosomiasis.

Prohibitin 1 (PHB1) controls growth and development and regulates proliferation and apoptosis in Schistosoma japonicum.

Schistosomiasis is a zoonotic parasitic disease caused by the trematode blood flukes of the genus Schistosoma. The prodigious egg output of females is the main cause of the disease in definitive hosts, while the female worm relies on continuous pairing with the male worm to fuel the growth and maturation of the reproductive organs and egg production. Prohibitin, which contains the functionally interdependent PHB1 and PHB2 subunits in human and some other species, has been proposed to participate in the cell proliferation and apoptosis regulation in mammals. However, little is known about the function of PHB homolog in the growth and reproductive development of schistosomes. Here, we reported the Phb1 gene that was structurally and evolutionarily conserved in Schistosoma japonicum when compared with that of other species from Caenorhabditis elegans to human. Real-time PCR detected that SjPhb1 was highly transcribed in the vitellaria of female worms. SjPhb1 knockdown achieved through the dsRNA-mediated RNAi in vivo resulted in retarded growth, decreased pairing, and fecundity in adult worms, as well as attenuated pathogenicity or virulence of worms to their hosts. Cell proliferation and apoptosis examination found decreased cell proliferation and increased cell apoptosis in SjPhb1 dsRNA-treated worms. Therefore, our study provides the first characterization of S. japonicum PHB1 and reveals its fundamental role in the regulation of growth and development of S. japonicum by specific dsRNA-mediated RNAi in vivo. Our findings prompt for a promising molecular of schistosomes that can be targeted to effectively retard the growth and development of the schistosomes.

Frequent upregulation of G9a promotes RelB-dependent proliferation and survival in multiple myeloma.

BACKGROUND: Multiple myeloma is an incurable hematological malignancy characterized by a heterogeneous genetic and epigenetic landscape. Although a number of genetic aberrations associated with myeloma pathogenesis, progression and prognosis have been well characterized, the role of many epigenetic aberrations in multiple myeloma remain elusive. G9a, a histone methyltransferase, has been found to promote disease progression, proliferation and metastasis via diverse mechanisms in several cancers. A role for G9a in multiple myeloma, however, has not been previously explored. METHODS: Expression levels of G9a/EHMT2 of multiple myeloma cell lines and control cells Peripheral Blood Mononuclear Cells (PBMCs) were analyzed. Correlation of G9a expression and overall survival of multiple myeloma patients were analyzed using patient sample database. To further study the function of G9a in multiple myeloma, G9a depleted multiple myeloma cells were built by lentiviral transduction, of which proliferation, colony formation assays as well as tumorigenesis were measured. RNA-seq of G9a depleted multiple myeloma with controls were performed to explore the downstream mechanism of G9a regulation in multiple myeloma. RESULTS: G9a is upregulated in a range of multiple myeloma cell lines. G9a expression portends poorer survival outcomes in a cohort of multiple myeloma patients. Depletion of G9a inhibited proliferation and tumorigenesis in multiple myeloma. RelB was significantly downregulated by G9a depletion or small molecule inhibition of G9a/GLP inhibitor UNC0642, inducing transcription of proapoptotic genes Bim and BMF. Rescuing RelB eliminated the inhibition in proliferation and tumorigenesis by G9a depletion. CONCLUSIONS: In this study, we demonstrated that G9a is upregulated in most multiple myeloma cell lines. Furthermore, G9a loss-of-function analysis provided evidence that G9a contributes to multiple myeloma cell survival and proliferation. This study found that G9a interacts with NF-κB pathway as a key regulator of RelB in multiple myeloma and regulates RelB-dependent multiple myeloma survival. G9a therefore is a promising therapeutic target for multiple myeloma.

Characteristics and function of cathepsin L3 from Schistosoma japonicum.

Schistosomiasis is still prevalent and seriously endangering the health of people and livestock in many countries. There have been great efforts to develop vaccines against schistosomiasis for prolonged protection in epidemic areas. Molecules from lung-stage schistosomula have been regarded as potential vaccine candidates against schistosomiasis. Our previous work has shown that cathepsin L3 from Schistosoma japonicum (SjCL3) is expressed in lung-stage schistosomula, but its role is not well known. In the present study, we characterized SjCL3 and detected its effect as a possible vaccine in vivo and in vitro. From the results of quantitative PCR (qPCR) and western blot, SjCL3 was present throughout the lifecycle of the worm, and its relative expressed level was higher in the liver eggs and adult worms than other stages. Additionally, immunofluorescence assay showed that SjCL3 was mainly concentrated in the eggshell, alimentary canal, and musculature of worms. Compared with the adjuvant group, the immunization of SjCL3 in mice resulted in a 28.9% decrease in worm burden and a 29.2% reduction in egg number in the host liver. In antibody-dependent cell-mediated cytotoxicity (ADCC) insecticidal experiments in vitro, the existence of SjCL3 could in part suppress adherence between macrophages and worm. The above results indicated that the immunization of SjCL3 could induce limited immune protection against S. japonicum infection in mice, and this protease played a role in breaking the process of ADCC, which was beneficial to the survival of worms.

Nanodiamond-Mediated Delivery of a G9a Inhibitor for Hepatocellular Carcinoma Therapy.

Hepatocellular carcinoma (HCC) is the most common primary liver cancer with high mortality but limited therapeutic options. Epigenetic regulations including DNA methylation and histone modification control gene expressions and play a crucial role during tumorigenesis. G9a, also known as EHMT2 (euchromatic histone-lysine N-methyltransferase 2), is a histone methyltransferase predominantly responsible for dimethylation of histone H3 lysine 9 (H3K9). G9a has been shown to play a key role in promoting tumor progression. Recent studies have identified that G9a is a critical mediator of HCC pathogenesis. UNC0646 is a G9a inhibitor that has shown potent in vitro efficacy. However, due to its water insolubility, the in vivo efficacy of UNC0646 is not satisfactory. In this study, nanodiamonds (NDs) were utilized as a drug delivery platform to improve in vivo delivery of this small-molecule inhibitor. Our results showed that ND-UNC0646 complexes could be rapidly synthesized by physical adsorption, meanwhile possessing favorable drug delivery properties and was able to improve the dispersibility of UNC0646 in water, therefore making it amenable for intravenous administration. The release profile of UNC0646 from ND-UNC0646 was demonstrated to be pH-responsive. Moreover, ND-UNC0646 maintained the biological functionality of UNC0646, with higher efficacy in reducing H3K9 methylation as well as enhanced invasion suppressive effects. Most importantly, increased in vivo efficacy was demonstrated using an orthotopic HCC mouse model, which paves the way of translating this small-molecule inhibitor toward HCC treatment. Our work demonstrates the potential of NDs in the clinical application for HCC treatment.

Stimuli-Responsive Nanodiamond-Based Biosensor for Enhanced Metastatic Tumor Site Detection.

Metastasis is often critical to cancer progression and linked to poor survival and drug resistance. Early detection of metastasis, as well as identification of metastatic tumor sites, can improve cancer patient survival. Thus, developing technology to improve the detection of cancer metastasis biomarkers can improve both diagnosis and treatment. In this study, we investigated the use of nanodiamonds to develop a stimuli-responsive metastasis detection complex that utilizes matrix metalloproteinase 9 (MMP9) as a metastasis biomarker, as MMP9 increased expression has been shown to be indicative of metastasis. The nanodiamond-MMP9 biosensor complex consists of nanodiamonds functionalized with MMP9-specific fluorescent-labeled substrate peptides. Using this design, protease activity of MMP9 can be accurately measured and correlated to MMP9 expression. The nanodiamond-MMP9 biosensor also demonstrated an enhanced ability to protect the base sensor peptide from nonspecific serum protease cleavage. This enhanced peptide stability, combined with a quantitative stimuli-responsive output function, provides strong evidence for the further development of a nanodiamond-MMP9 biosensor for metastasis site detection. More importantly, this work provides the foundation for use of nanodiamonds as a platform for stimuli-responsive biosensors and theranostic complexes that can be implemented across a wide range of biomedical applications.