CircTHBS1 promotes trophoblast cell migration and invasion and inhibits trophoblast apoptosis by regulating miR-136-3p/IGF2R axis.

The precise molecular mechanism behind fetal growth restriction (FGR) is still unclear, although there is a strong connection between placental dysfunction, inadequate trophoblast invasion, and its etiology and pathogenesis. As a new type of non-coding RNA, circRNA has been shown to play a crucial role in the development of FGR. This investigation identified the downregulation of hsa_circ_0034533 (circTHBS1) in FGR placentas through high-sequencing analysis and confirmed this finding in 25 clinical placenta samples using qRT-PCR. Subsequent in vitro functional assays demonstrated that silencing circTHBS1 inhibited trophoblast proliferation, migration, invasion, and epithelial mesenchymal transition (EMT) progression and promoted apoptosis. Furthermore, when circTHBS1 was overexpressed, cell function experiments showed the opposite result. Analysis using fluorescence in situ hybridization revealed that circTHBS1 was primarily found in the cytoplasmic region. Through bioinformatics analysis, we anticipated the involvement of miR-136-3p and IGF2R in downstream processes, which was subsequently validated through qRT-PCR and dual-luciferase assays. Moreover, the inhibition of miR-136-3p or the overexpression of IGF2R partially reinstated proliferation, migration, and invasion abilities following the silencing of circTHBS1. In summary, the circTHBS1/miR-136-3p/IGF2R axis plays a crucial role in the progression and development of FGR, offering potential avenues for the exploration of biological indicators and treatment targets.

CO-Tolerant Hydrogen Oxidation Electrocatalysts for Low-Temperature Hydrogen Fuel Cells.

The severe performance degradation of low-temperature hydrogen fuel cells upon exposure to trace amounts of carbon monoxide (CO) impurities in reformate hydrogen fuels is one of the challenges that hinders their commercialization. Despite significant efforts that have been made, the CO-tolerance performance of electrocatalysts for the hydrogen oxidation reaction (HOR) is still unsatisfactory. This Perspective discusses the path forward for the rational design of CO-tolerant HOR electrocatalysts. The fundamentals of the CO-tolerant mechanisms on commercialized platinum group metal (PGM) electrocatalysts via either promoting CO electrooxidation or weakening CO adsorption are provided, and comprehensive discussions based on these strategies are presented with typical examples. Given the recent progress, some emerging strategies, including blocking CO diffusion with a barrier layer and developing non-PGM HOR catalysts, are also discussed. We conclude with a discussion of the strengths and limitations of these strategies along with the perspectives of the major challenges and opportunities for future research on CO-tolerant HOR electrocatalysts.

A large deletion in TSC2 causes tuberous sclerosis complex by dysregulating PI3K/AKT/mTOR signaling pathway.

BACKGROUND/AIM: Tuberous sclerosis complex (TSC) is a multi-system syndrome caused by loss-of-function mutation in TSC1 or TSC2. Most TSC patients present with cardiac rhabdomyoma or cortical tubers during fetal life, and the symptoms are not uniform as their age. The gene products of TSC1/2 are components of the TSC protein complex and are important role in the PI3K/AKT/mTOR (PAM) signaling pathway. Based on three members of a family with variable expressivity, the purpose of this study was to clarify the clinical features of TSC in different age groups and to analyze the genetic characteristics of TSC2 gene. METHODS: Clinical exome sequencing and co-segregation were used to identify a three-generation family with four affected individuals. HEK-293T cell model was constructed for subsequent experiments. Quantitative RT-PCR, western blotting, and subcellular localization were used to analyze the expression effect of TSC2 mutation. CCK-8 assay, wound healing assay, and cell cycle analysis were used to analyze the function effect of TSC2 mutation. RESULT: We identified a TSC family with heterozygous deletion of exon 4 in TSC2 by clinical exon sequencing. Sanger sequencing indicated that the affected individuals have 2541-bp deletion that encompassed exon 4 and adjacent introns. Deletion of exon 4 decreased the TSC2 mRNA and protein levels in HEK-293T cells, and activated the PI3K/AKT/mTOR pathway, thereby altering the cell cycle and promoting cell proliferation and migration. CONCLUSION: We confirmed the pathogenicity of the large deletion in TSC2 in a three- generations family.. Deletion of exon 4 of TSC2 affected cell proliferation, migration, and cell cycle via abnormal activation of the PAM pathway. This study evaluated the pathogenic effect of deletion of exon 4 of TSC2 and investigated the underlying mechanism.

Ru-Doped Ultrasmall Cu Nanoparticles Decorated with Carbon for Electroreduction of Nitrate to Ammonia.

Electrocatalytic nitrate reduction reaction offers a sustainable approach to treating wastewater and synthesizing high-value ammonia under ambient conditions. However, electrocatalysts with low faradaic efficiency and selectivity severely hinder the development of nitrate-to-ammonia conversion. Herein, Ru-doped ultrasmall copper nanoparticles loaded on a carbon substrate (Cu-Ru@C) were fabricated by the pyrolysis of Cu-BTC metal-organic frameworks (MOFs). The Cu-Ru@C-0.5 catalyst exhibits a high faradaic efficiency (FE) of 90.4% at -0.6 V (vs RHE) and an ammonia yield rate of 1700.36 µg h-1mgcat.-1 at -0.9 V (vs RHE). Moreover, the nitrate conversion rate is almost 100% over varied pHs (including acid, neutral, and alkaline electrolytes) and different nitrate concentrations. The remarkable performance is attributed to the synergistic effect between Cu and Ru and the excellent conductivity of the carbon substrate. This work will open an exciting avenue to exploring MOF derivatives for ambient ammonia synthesis via selective electrocatalytic nitrate reduction.

Structure and Defect Engineering Synergistically Boost High Solar-to-Chemical Conversion Efficiency of Cerium oxide/Au Hollow Nanomushrooms for Nitrogen Photofixation.

Photocatalytic nitrogen fixation using solar illumination under ambient conditions is a promising strategy for production of the indispensable chemical NH3 . However, due to the catalyst's limitations in solar energy utilization, loss of hot electrons during transfer, and low nitrogen adsorption and activation capacity, the unsatisfactory solar-to-chemical conversion (SCC) efficiencies of most photocatalysts limit their practical applications. Herein, cerium oxide nanosheets with abundant strain-VO defects were anchored on Au hollow nanomushroom through atomically sharp interfaces to construct a novel semiconductor/plasmonic metal hollow nanomushroom-like heterostructure (denoted cerium oxide-AD/Au). Plasmonic Au extended the absorption of light from the visible to the second near-infrared region. The superior interface greatly enhanced the transfer efficiency of hot electrons. Abundant strain-VO defects induced by interfacial compressive strain promoted adsorption and in situ activation of nitrogen, and such synergistic promotion of strain and VO defects was further confirmed by density functional theory calculations. The judicious structural and defect engineering co-promoted the efficient nitrogen photofixation of the cerium oxide-AD/Au heterostructures with a SCC efficiency of 0.1 % under simulated AM 1.5G solar illumination, which is comparable to the average solar-to-biomass conversion efficiency of natural photosynthesis by typical plants, thus exhibiting significant potential as a new candidate for artificial photosynthesis.

Dysfunction of ZNF554 promotes ROS-induced apoptosis and autophagy in Fetal Growth Restriction via the p62-Keap1-Nrf2 pathway.

Fetal growth restriction (FGR) is one of the most common complications of an abnormal pregnancy. Placental dysplasia has been established as a significant contributing factor to FGR. Zinc finger protein 554 (ZNF554) is a member of the Krüppel-associated box domain zinc finger protein subfamily, primarily expressed in the placenta and essential for maintaining normal pregnancy outcomes. However, its precise role in FGR remains uncertain. In this study, we confirmed that ZNF554 was low expressed in the placenta of the FGR pregnancy. To further elucidate the impact of ZNF554 on trophoblasts, we conducted experiments using siRNA and overexpression plasmids on HTR8/SVneo and JEG3 cells. Our findings revealed that silencing ZNF554 increased apoptosis and inhibited migration and invasion, while overexpression reduced apoptosis and promoted migration and invasion. Notably, ZNF554 knockdown decreased cellular antioxidant capacity and elevated the production of reactive oxygen species (ROS). Conversely, ZNF554 activated the nuclear factor E2-related factor 2 (NRF2) signaling pathway, exerting its antioxidant effects. Additionally, ZNF554 knockdown promoted cellular autophagy by suppressing P62 and enhancing LC3-II/LC3-I expression. Importantly, the antioxidant N-acetylcysteine (NAC) partially mitigated the impact of ZNF554 knockdown on mitochondrial ROS in trophoblast cells and subsequent effects on cellular autophagy and apoptosis. In conclusion, our results suggest that ZNF554 plays a pivotal role in modulating trophoblast cell invasion and may serve as a prognostic marker and potential therapeutic target for FGR.

Powerful Orbital Hybridization of Copper-Silver Bimetallic Nanosheets for Electrocatalytic Nitrogen Reduction to Ammonia.

Electrochemical nitrogen reduction (eNRR) is a promising strategy to replace the energy- and capital-intensive Haber-Bosch process. Unfortunately, the low selectivity of the eNRR process impedes the industrial application of this approach. In this work, a highly efficient and stable NRR electrocatalyst is obtained via coreduction of Cu and Ag precursors using the holly leaves as reducing agents. The as-obtained Cu3Ag bimetallic nanosheets exhibit excellent NRR performance with an NH3 production rate of 31.3 µg h-1 mg-1cat. and a Faradaic efficiency of 31.3% at -0.2 V vs RHE. According to density functional theory (DFT) calculation, the outstanding performance of Cu3Ag bimetallic nanosheets could be caused by the fact that Ag optimizes the 3d orbital occupation of Cu and synergistically enhances the charge transfer during the NRR process, resulting in a suitable adsorption strength of the intermediates.

Selenite-Decorated Polycrystalline NiO Nanosheets Generated from Cathodic Reconstruction for Electrocatalytic Hydrogen Production.

Precatalyst reconstruction in alkaline hydrogen evolution reaction (HER) usually leads to changes in the morphology, composition, and structure, thus improving the catalytic activity, which recently receives intensive attention. However, the design strategies of cathodic reconstruction and the structural features of reconstruction products have not achieved a profound understanding. Here, from the point of thermodynamic stability, metastable nickel selenite dihydrate (NiSeO3·2H2O) is deliberately fabricated as a precatalyst to comprehensively study the reconstruction dynamics in alkaline HER. Multiple in/ex situ techniques capture the geometric, component, and phase evolutions, proving that NiSeO3·2H2O can be transformed into SeO32--decorated polycrystalline NiO nanosheets with rich active sites and good conductivity under alkaline HER conditions, which act as a real catalytic active species. Density functional theory calculations demonstrate that the adsorption of SeO32- can further promote the HER activity of NiO due to the optimized free energy of water activation and hydrogen adsorption. As a result, the SeO32--NiO catalyst exhibits a low overpotential at -10 mA cm-2 (90 mV) and long-term stability (>100 h). This work highlights the targeted design of precatalyst to trigger and utilize cathodic reconstruction and provides an available method for the development of adsorption-modulated efficient electrocatalysts.

Increased Oxygen Vacancies in CeO2 for Improved Electrocatalytic Nitrogen Reduction Performance.

Electrochemical nitrogen fixation is a sustainable and economical strategy to produce ammonia. However, fabricating efficient electrocatalysts for nitrogen fixation is still challenging. Theoretical predictions prove that the oxygen vacancy is able to modulate the electronic state of CeO2 and enhance its electrical conductivity, thus promoting the electrochemical nitrogen reduction reaction (NRR) process. Herein, CeO2 with high oxygen vacancy concentration was prepared via a two-step pyrolysis strategy of Ce metal-organic frameworks (MOFs, denoted H-CeO2). Compared to CeO2 with low oxygen vacancy concentration synthesized via one-step pyrolysis of Ce-MOFs (denoted L-CeO2), H-CeO2 exhibits a large NH3 yield rate (25.64 µg h-1 mgcat-1 at -0.5 V vs reversible hydrogen electrode, RHE) and high faradaic efficiency (FE, 6.3% at -0.4 V vs RHE).

Performance of whole-genome promoter nucleosome profiling of maternal plasma cell-free DNA for prenatal noninvasive prediction of fetal macrosomia: a retrospective nested case-control study in mainland China.

BACKGROUND: Fetal macrosomia is common occurrence in pregnancy, which is associated with several adverse prognosis both of maternal and neonatal. While, the accuracy of prediction of fetal macrosomia is poor. The aim of this study was to develop a reliable noninvasive prediction classifier of fetal macrosomia. METHODS: A total of 3600 samples of routine noninvasive prenatal testing (NIPT) data at 12+ 0-27+ 6 weeks of gestation, which were subjected to low-coverage whole-genome sequencing of maternal plasma cell-free DNA (cfDNA), were collected from three independent hospitals. We identified set of genes with significant differential coverages by comparing the promoter profiling between macrosomia cases and controls. We selected genes to develop classifier for noninvasive predicting, by using support vector machine (SVM) and logistic regression models, respectively. The performance of each classifier was evaluated by area under the curve (AUC) analysis. RESULTS: According to the available follow-up results, 162 fetal macrosomia pregnancies and 648 matched controls were included. A total of 1086 genes with significantly differential promoter profiling were found between pregnancies with macrosomia and controls (p < 0.05). With the AUC as a reference，the classifier based on SVM (CMA-A2) had the best performance, with an AUC of 0.8256 (95% CI: 0.7927-0.8586). CONCLUSIONS: Our study provides that assessing the risk of fetal macrosomia by whole-genome promoter nucleosome profiling of maternal plasma cfDNA based on low-coverage next-generation sequencing is feasible.

MOF-Derived Cu3P nanoparticles coated with N-doped carbon for nitrogen fixation.

Electrochemical nitrogen reduction is a significant alternative route for synthesizing ammonia, but constructing efficient catalysts for electrochemical nitrogen fixation still faces tough challenges. In this work, Cu3P@NC (NC: nitrogen-doped carbon) nanosheets were prepared via the low-temperature pyrolysis-phosphating of Cu-MOFs. When applied to the nitrogen reduction reaction, Cu3P@NC exhibited a high ammonia yield rate of 10.4 µg h-1 mg-1cat at -0.3 V (vs. RHE) and a faradaic efficiency (FE) of 6.3% at -0.1 V (vs. RHE). The outstanding performance was attributed to the large electrochemical surface area and the defects induced as a result of N doping, which helped enhance N2 adsorption. This work provides a novel strategy for preparing N-doped carbon materials for wide-ranging applications.

Amorphous ReS2 decorated TiO2 nanowire arrays for highly-efficient nitrogen reduction.

Herein, we constructed amorphous ReS2 nanosheets anchored on TiO2 nanowires by delicately forming Ti-O-Re bonds, which exhibited considerable average ammonia yield and faradaic efficiency as high as 5.3 µg h-1 cm-2cat. and 49.8% at an applied potential of -0.2 V (vs. RHE) in 0.1 M Na2SO4. The high NRR performance could be attributed to the amorphous feature of the ReS2 nanosheets, the rich oxygen vacancies in the TiO2 nanowires and their semiconducting feature, which not only facilitate the sufficient exposure of active sites but also efficiently boost the faradaic efficiency due to the good regulation of surface proton or electron accessibility.

Complete Genome Sequence of the Red-Pigmented Strain Serratia marcescens SCQ1 and Its Four Spontaneous Pigment Mutants.

Serratia marcescens SCQ1 is a red-pigmented bacterium isolated from silkworm larva with septicemia. Pigment-deficient spontaneous mutants arise when S. marcescens SCQ1 is incubated under relatively stable laboratory conditions for a long time. Here, we present the complete genome sequence of SCQ1 and the resequenced genomes of four spontaneous pigment mutants.

In vitro transcriptomes analysis identifies some special genes involved in pathogenicity difference of the Beauveria bassiana against different insect hosts.

Typical entomopathogenic filamentous fungi such as Beauveria bassiana infect susceptible hosts via penetration of insect cuticle. The pathogenicity of B. bassiana strain to diverse insect hosts is different. While the molecular mechanisms of B. bassiana adapt to different insects are not well clear. B. bassiana GXsk1011 is a hyper-virulent strain from silkworm, which was investigated on the metabolic responses to three cuticle extracts of Bombyx mori, Helicoverpa armigera and Clanis bilineata at 24 h by RNA-seq method. A total of 638 up- and 400 down-regulated differentially expressed genes (DEGs) were identified in B. bassiana grown on H. armigera compared with B. mori, and 910 up- and 401 down-regulated genes for C. bilineata compared with B. mori. Functional categorization showed that DEGs are mainly involved in metabolic processes, localization, catalytic activity and transporter activity. Analysis of 20 highest fold change genes in DEGs showed that when B. bassiana transferred to non-original hosts as H. armigera and C. bilineata, the adhesion (Mad1), protease (Pr2) and cell surface protein (BBA_09174), etc. were down-regulated. While the class III chitinase ChiA2 (BBA_05353, Bbchi-17), major allergen Asp f 2-like protein (BBA_05395, Bb-f2) and nonribosomal peptide synthase, etc. were up-regulated. The secretory lipase that responded to H. armigera and the phosphate permease responded to C. bilineata were also up-regulated in the Top 20 DEGs. These special expressed genes indicate when the B. bassiana transferred to non-original hosts (or called as non-natural hosts), the strain appeared the changes of metabolic response and infection strategies to adapt to new hosts, and implied the key actions of infected adaptation were to break the barrier of different cuticle chitin component and against the immune stress of hosts. This study provided an insight into the B. bassiana that with wide host ranges how to adapt to infect different insect hosts, which will help us to further understand the pathogenesis of B. bassiana infection.

Bonding interface boosts the intrinsic activity and durability of NiSe@Fe2O3 heterogeneous electrocatalyst for water oxidation.

The intrinsic activity and durability of oxygen evolution reaction (OER) electrocatalysts are mainly dominated by the surface and interface properties of active materials. Herein, a core-shell heterogeneous structure (NF/NiSe@Fe2O3) is fabricated via two-step hydrothermal method, which exhibits a low overpotential of 220 mV (or 282 mV) at 10 mA/cm2 (or 200 mA/cm2), a small Tafel slope of 36.9 mV/dec, and long-term stability (~230 h) in 1 mol/L KOH for OER. X-ray photoelectron spectroscopy and X-ray absorption spectroscopy reveal the (oxy)hydroxide-rich surface and strong coupling interface between NiSe and Fe2O3 via the Fe-Se bond. Density functional theory calculation suggests that the d-band center and electronic state of NiSe@Fe2O3 heterojunction are well optimized due to the formation of Fe-Se bond, which is favorable for the enhanced OER activity because of the easy adsorption of oxygen-containing intermediates and desorption of O2 in the OER process. In addition, the unique core-shell structure and robust bonding interface are responsible for the good stability for OER. This work provides fundamental insights on the bonding effect that determine the performance of OER electrocatalyst.

A Novel Mutation in ATRX Causes Alpha-Thalassemia X-Linked Intellectual Disability Syndrome in a Han Chinese Family.

OBJECTIVE: To analyze genetic mutations in a Chinese pedigree affected with Alpha-thalassemia X-linked intellectual disability syndrome, providing a precise diagnosis and genetic counseling. METHODS: Clinical data was collected. A novel alternative splicing variant detected by whole-exome sequencing was validated by Sanger sequencing. The functional effect of the mutation was predicted with Mutation Tasting. The analysis of 5' splice site score was estimated with MaxEntScan. Changes in amino acid sequencing were predicted with Mutalyzer. The tertiary structures of the wild type and mutation-carrying protein were predicted by I-TASSER. RNA was extracted from peripheral blood lymphocytes from the proband, his mother and a healthy control. Quantitative Real-Time PCR was used to detect mRNA expression. RESULTS: The proband presented with severe intellectual disability, developmental delay, characteristic facies, seizures and cryptorchidism. A novel hemizygous duplication mutation in the ATRX gene in a splice site between exons 3 and 4, NM_000489: c.189+1dupG, was identified with WES in the proband. Sanger sequencing confirmed that the mutation was inherited from his mother, who carried a heterozygous mutation, while his father was not affected. Bioinformatics analysis indicated that the splicing region where the mutation was located is highly conserved and the variant was damaging, producing a truncated protein due to the premature translation of a stop codon. Sanger sequencing with the Quantitative Real-Time PCR product containing a G base inserted between bases 189 and 190. The level of mRNA expression showed that ATRX gene transcription decreased due to the mutation (P < 0.05). CONCLUSIONS: A novel mutation in ATRX was found in this pedigree and was confirmed to be pathogenic through functional studies. Our research expanded the spectrum of ATRX gene mutations, providing a precise diagnosis and a basis for genetic counseling.

Transcriptomic Analysis Reveals Competitive Growth Advantage of Non-pigmented Serratia marcescens Mutants.

Serratia marcescens is a common bacterium well-known for the red secondary metabolite prodigiosin. However, color mutants have long been described. Non-pigmented strains can be found to exist both naturally and under laboratory conditions. It is unclear why S. marcescens loses prodigiosin synthesis capacity in certain conditions. In the present study, we find that the spontaneous color mutants arise within a few generations (about five passages) and rapidly replace the wild-type parent cells (about 24 passages), which indicates a growth advantage of the former. Although, the loss of prodigiosin synthesis genes (pigA-N) is frequently reported as the major reason for pigment deficiency, it was unexpected that the whole gene cluster is completely preserved in the different color morphotypes. Comparative transcriptomic analysis indicates a dramatic variation at the transcriptional level. Most of the pig genes are significantly downregulated in the color morphotypes which directly lead to prodigiosin dyssynthesis. Besides, the transcriptional changes of several other genes have been noticed, of which transcriptional regulators, membrane proteins, and nearly all type VI secretion system (T6SS) components are generally downregulated, while both amino acid metabolite and transport systems are activated. In addition, we delete the transcription regulator slyA to generate a non-pigmented mutant. The ΔslyA strain loses prodigiosin synthesis capacity, but has a higher cell density, and surprisingly enhances the virulence as an entomopathogen. These data indicate that S. marcescens shuts down several high-cost systems and activates the amino acid degradation and transport pathways at the transcriptional level to obtain extra resources, which provides new insights into the competitive growth advantage of bacterial spontaneous color mutants.

Whole-Genome Promoter Profiling of Plasma DNA Exhibits Diagnostic Value for Placenta-Origin Pregnancy Complications.

Placenta-origin pregnancy complications, including preeclampsia (PE), gestational diabetes mellitus (GDM), fetal growth restriction (FGR), and macrosomia (MA) are common occurrences in pregnancy, resulting in significant morbidity and mortality for both mother and fetus. However, despite their frequency, there are no reliable methods for the early diagnosis of these complications. Since cfDNA is mainly derived from placental trophoblasts and maternal hematopoietic cells, it might have information for gene expression which can be used for disease prediction. Here, low coverage whole-genome sequencing on plasma DNA from 2,199 pregnancies is performed based on retrospective cohorts of 3,200 pregnant women. Read depth in the promoter regions is examined to define read-depth distribution patterns of promoters for pregnancy complications and controls. Using machine learning methods, classifiers for predicting pregnancy complications are developed. Using these classifiers, complications are successfully predicted with an accuracy of 80.3%, 78.9%, 72.1%, and 83.0% for MA, FGR, GDM, and PE, respectively. The findings suggest that promoter profiling of cfDNA may be used as a biological biomarker for predicting pregnancy complications at early gestational age.

Engineering the valence state of ZIF-67 by Cu2O for efficient nonenzymatic glucose detection.

The valence state regulation of Co-based electrocatalysts is extremely important and greatly challenging to enhance the electrochemical performance toward glucose oxidation. Herein, Cu2O@ZIF-67 composites with fine-tuned valence states were rationally constructed for boosting glucose oxidation. X-ray photoelectron spectroscopy (XPS) and cyclic voltammetry (CV) analysis confirm that the content of the high valence state Co (Co3+) in Cu2O@ZIF-67 is much higher than that in the individual ZIF-67 due to the synergistic effects between ZIF-67 and Cu2O. As a result, Co3+-rich Cu2O@ZIF-67 composite exhibits remarkable activity toward glucose electro-oxidation with two linear response ranges of glucose concentration, from 0.01 to 10 mM and 10 to 16.3 mM, high sensitivities of the linear ranges (307.02 and 181.34 µA mM-1 cm-2) as well as a low detection limit (6.5 µM). This research provides a novel avenue for the progress of highly efficient electrocatalysts for nonenzymatic glucose oxidation.

A novel MoNi@Ni(OH)2 heterostructure with Pt-like and stable electrocatalytic activity for the hydrogen evolution reaction.

A novel Mo0.84Ni0.16@Ni(OH)2 heterostructure was successfully fabricated. Owing to the unique interface structure and strong synergistic effects between different components, the heterostructure shows enhanced activity for the hydrogen evolution reaction (HER), outperforming that of the state-of-the-art Pt/C catalyst.