Glycerophosphodiesters inhibit lysosomal phospholipid catabolism in Batten disease.

Batten disease, the most prevalent form of neurodegeneration in children, is caused by mutations in the CLN3 gene, which encodes a lysosomal transmembrane protein. CLN3 loss leads to significant accumulation of glycerophosphodiesters (GPDs), the end products of glycerophospholipid catabolism in the lysosome. Despite GPD storage being robustly observed upon CLN3 loss, the role of GPDs in neuropathology remains unclear. Here, we demonstrate that GPDs act as potent inhibitors of glycerophospholipid catabolism in the lysosome using human cell lines and mouse models. Mechanistically, GPDs bind and competitively inhibit the lysosomal phospholipases PLA2G15 and PLBD2, which we establish to possess phospholipase B activity. GPDs effectively inhibit the rate-limiting lysophospholipase activity of these phospholipases. Consistently, lysosomes of CLN3-deficient cells and tissues accumulate toxic lysophospholipids. Our work establishes that the storage material in Batten disease directly disrupts lysosomal lipid homeostasis, suggesting GPD clearance as a potential therapeutic approach to this fatal disease.

Extracellular production of a thermostable Cellvibrio endolytic ß-agarase in Escherichia coli for agarose liquefaction.

Four GH16 family ß-agarases (GH16A, GH16B, GH16C, and GH16D), originated from an agarolytic bacterium Cellvibrio sp. KY-GH-1, were expressed in an Escherichia coli system and their activities were compared. Only GH16B (597 amino acids, 63.8 kDa), with N-terminal 22-amino acid signal sequence, was secreted into the culture supernatant and demonstrated a robust endolytic agarose hydrolyzing activity for producing neoagarotetraose (NA4) and neoagarohexaose (NA6) as end products. The optimal temperature and pH for the enzyme activity were 50 °C and 7.0, respectively. The enzyme was stable up to 50 °C and over a pH range of 5.0-8.0. The kinetic parameters, including Km, Vmax, kcat, and kcat/Km, of GH16B ß-agarases for agarose were 14.40 mg/mL, 542.0 U/mg, 576.3 s-1, and 4.80 × 106 s-1 M-1, respectively. The addition of 1 mM MnCl2 and 15 mM tris(2-carboxyethyl)phosphine enhanced the enzymatic activity. When agarose or neoagaro-oligosaccharides were used as substrates, the end products of enzymatic catalysis were NA4 and NA6, whereas agaropentaose was produced along with NA4 and NA6 when agaro-oligosaccharides were used as substrates. Treatment of 9%[w/v] melted agarose with the enzyme (1.6 µg/mL) under continuous magnetic stirring at 50 °C for 14 h resulted in efficient agarose liquefaction into NA4 and NA6. Purification of NA4 and NA6 from the enzymatic hydrolysate (9%[w/v] agarose, 20 mL) via Sephadex G-15 column chromatography yielded ~ 650 mg NA4/~ 900 mg NA6 (i.e., ~ 85.3% of the theoretical maximum yield). These findings suggest that the recombinant thermostable GH16B ß-agarase is useful for agarose liquefaction to produce NA4 and NA6.

The impact of containment measures and monetary and fiscal responses on US financial markets during the COVID-19 pandemic.

This paper analyses the effects of containment measures and monetary and fiscal responses on US financial markets during the Covid-19 pandemic. More specifically, it applies fractional integration methods to analyse their impact on the daily S&P500, the US Treasury Bond Index (USTB), the S&P Green Bond Index (GREEN) and the Dow Jones (DJ) Islamic World Market Index (ISLAM) over the period 1/01/2020-10/03/2021. The results suggest that all four indices are highly persistent and exhibit orders of integration close to 1. A small degree of mean reversion is observed only for the S&P500 under the assumption of white noise errors and USTB with autocorrelated errors; therefore, market efficiency appears to hold in most cases. The mortality rate, surprisingly, seems to have affected stock and bond prices positively with autocorrelated errors. As for the policy responses, both the containment and fiscal measures had a rather limited impact, whilst there were significant announcement effects which lifted markets, especially in the case of monetary announcements. There is also evidence of a significant, positive response to changes in the effective Federal funds rate, which suggests that the financial industry, mainly benefiting from interest rises, plays a dominant role.

Density of imaginary multiplicative chaos via Malliavin calculus.

We consider the imaginary Gaussian multiplicative chaos, i.e. the complex Wick exponential µ ß : = : e i ß Γ ( x ) : for a log-correlated Gaussian field Γ in d ≥ 1 dimensions. We prove a basic density result, showing that for any nonzero continuous test function f, the complex-valued random variable µ ß ( f ) has a smooth density w.r.t. the Lebesgue measure on C . As a corollary, we deduce that the negative moments of imaginary chaos on the unit circle do not correspond to the analytic continuation of the Fyodorov-Bouchaud formula, even when well-defined. Somewhat surprisingly, basic density results are not easy to prove for imaginary chaos and one of the main contributions of the article is introducing Malliavin calculus to the study of (complex) multiplicative chaos. To apply Malliavin calculus to imaginary chaos, we develop a new decomposition theorem for non-degenerate log-correlated fields via a small detour to operator theory, and obtain small ball probabilities for Sobolev norms of imaginary chaos.

The process of transferring negative impulses in capital markets - a wavelet analysis.

The empirical research that is presented herein deals with the process of transferring negative impulses in capital markets during the subprime crisis (contagion, comovements, crisis transmission and shocks). A significant and positive contribution of the research conducted is the demonstration of how the wavelet analysis can be used in examining the various responses of the financial markets. The first stage of the research involved an analysis of the response of seven European markets (CAC40, DAX, FTSE100, IBEX, ATHEX, BUX and WIG20 indexes) to the proceedings in the US market, exemplified by the Dow Jones Industrial Average Index. The second stage involved examining the relationships of strong European markets (CAC40, DAX, FTSE100), and then the impact that the strongest German market DAX had on four other and weaker European markets - two from Western Europe (IBEX, ATHEX) and two from Central-Eastern Europe (BUX and WIG20). This article presents a methodological approach to transfer impulses on capital markets.

Whole-genome analysis of a rare G15P[21] group A rotavirus detected at a dairy cattle farm.

Group A rotaviruses (RVAs) are a major cause of severe enteritis in humans and animals. RVAs have been identified in several animal species and their genetic diversity, the segmented nature of their RNA genome and the ability to spill over from one species to another can generate new RVA strains. In this study, we investigated the genome constellations of an unusual, rare, bovine RVA strain, G15P[21], identified from a farm with neonatal diarrhoea of calves in 2006. In parallel, the genome constellations of other RVA strains with different G/P types identified from the same farm in the same time span (2006-2008) were analysed. The genome constellation of strain K53 was G15-P[21]-I2-R2-C2-M2-A13-N2-T9-E2-H3 and was similar, overall, to that of the other bovine RVA strains (G6/10-P[11]-I2-R2-C2-M2-A13-N2-T6-E2-H3) with the exception of the NSP3 segment (T9 vs T6). This study describes RVA genomes with different genotype combinations isolated at a farm and also contributes to the understanding of the diversity and evaluation of rotavirus in a global context.

Operon dynamics with state dependent transcription and/or translation delays.

Transcription and translation retrieve and operationalize gene encoded information in cells. These processes are not instantaneous and incur significant delays. In this paper we study Goodwin models of both inducible and repressible operons with state-dependent delays. The paper provides justification and derivation of the model, detailed analysis of the appropriate setting of the corresponding dynamical system, and extensive numerical analysis of its dynamics. Comparison with constant delay models shows significant differences in dynamics that include existence of stable periodic orbits in inducible systems and multistability in repressible systems. A combination of parameter space exploration, numerics, analysis of steady state linearization and bifurcation theory indicates the likely presence of Shilnikov-type homoclinic bifurcations in the repressible operon model.

Saikosaponin-d alleviates hepatic fibrosis through regulating GPER1/autophagy signaling.

BACKGROUND: Hepatic fibrosis is the final pathway of chronic liver disease characterized by excessive accumulation of extracellular matrix (ECM), which eventually develop into cirrhosis and liver cancer. Emerging studies demonstrated that Saikosaponin-d (SSd) exhibits a protective role in liver fibrosis. However, the mechanism underlying anti-liver fibrosis of SSd in vivo and in vitro remains unclear. METHODS AND RESULTS: Transforming growth factor (TGF)-ß and carbon tetrachloride (CCl4) were used for creating liver fibrosis model in vitro and in vivo, respectively. The role of SSd in regulating liver fibrosis was assessed through Sirius red and Masson staining, and IHC assay. We found that SSd attenuated remarkably CCl4-induced liver fibrosis as evidenced by decreased collagen level, and decreased expression of fibrotic markers Col 1 and α-SMA. Meanwhile, SSd repressed autophagy activation as suggested by decreased BECN1 expression and increased p62 expression. Compared with HSCs from CCl4-treated group, the primary HSCs from SSd-treated mice exhibited a marked inactivation of autophagy. Mechanistically, SSd treatment enhanced the expression of GPER1 in primary HSCs and in TGF-ß-treated LX-2 cells. GPER1 agonist G1 repressed autophagy activation, whereas GPER1 antagonist G15 activated autophagy and G15 also damaged the function of SSd on suppressing autophagy, leading to subsequent increased levels of fibrotic marker level in LX-2 cells. CONCLUSIONS: Our findings highlight that SSd alleviates hepatic fibrosis by regulating GPER1/autophagy pathway.

Pepper root rot resistance and pepper yield are enhanced through biological agent G15 soil amelioration.

Pepper root rot is a serious soil-borne disease that hinders pepper production, and efforts are being made to identify biological agents that can prevent and control pepper root rot. Our group recently discovered and produced a biological agent, named G15, which reduces the diversity and richness of fungi and bacteria when applied to pepper fields. In the soil of the G15-treatment condition, the pathogenic fungus Fusarium was inhibited, while the richness of beneficial bacteria Rhodanobacter was increased. Also, the ammonia nitrogen level was decreased in the G15-treatment soil, and the pH, total carbon, and total potassium levels were increased. Compared to the control condition, pepper yield was increased in the treatment group (by 16,680 kg acre-1). We found that G15 could alter the microbial community structure of the pepper rhizosphere. These changes alter the physical and chemical properties of the soil and, ultimately, improve resistance to pepper root rot and increase pepper yield.

Estrogen-dependent KCa1.1 modulation is essential for retaining neuroexcitation of female-specific subpopulation of myelinated Ah-type baroreceptor neurons in rats.

Female-specific subpopulation of myelinated Ah-type baroreceptor neurons (BRNs) in nodose ganglia is the neuroanatomical base of sexual-dimorphic autonomic control of blood pressure regulation, and KCa1.1 is a key player in modulating the neuroexcitation in nodose ganglia. In this study we investigated the exact mechanisms underlying KCa1.1-mediated neuroexcitation of myelinated Ah-type BRNs in the presence or absence of estrogen. BRNs were isolated from adult ovary intact (OVI) or ovariectomized (OVX) female rats, and identified electrophysiologically and fluorescently. Action potential (AP) and potassium currents were recorded using whole-cell recording. Consistently, myelinated Ah-type BRNs displayed a characteristic discharge pattern and significantly reduced excitability after OVX with narrowed AP duration and faster repolarization largely due to an upregulated iberiotoxin (IbTX)-sensitive component; the changes in AP waveform and repetitive discharge of Ah-types from OVX female rats were reversed by G1 (a selective agonist for estrogen membrane receptor GPR30, 100 nM) and/or IbTX (100 nM). In addition, the effect of G1 on repetitive discharge could be completely blocked by G15 (a selective antagonist for estrogen membrane receptor GPR30, 3 µM). These data suggest that estrogen deficiency by removing ovaries upregulates KCa1.1 channel protein in Ah-type BRNs, and subsequently increases AP repolarization and blunts neuroexcitation through estrogen membrane receptor signaling. Intriguingly, this upregulated KCa1.1 predicted electrophysiologically was confirmed by increased mean fluorescent intensity that was abolished by estrogen treatment. These electrophysiological findings combined with immunostaining and pharmacological manipulations reveal the crucial role of KCa1.1 in modulation of neuroexcitation especially in female-specific subpopulation of myelinated Ah-type BRNs and extend our current understanding of sexual dimorphism of neurocontrol of BP regulation.

Anti-inflammatory and Antioxidant Activity of Peptides From Ethanol-Soluble Hydrolysates of Sturgeon (Acipenser schrenckii) Cartilage.

Research has shown that cartilage containing chondroitin sulfate and protein presents versatile bioactivities. Chondroitin sulfate in cartilage is beneficial to activate the immune system while the protein/peptide has not been fully understood. The current study investigated the antioxidant and anti-inflammatory properties of ethanol-soluble hydrolysates of sturgeon cartilage (ESCH) prepared through hot-pressure, enzymatic hydrolysis and ethanol extraction. UV spectrum, IR and agarose gel electrophoresis results suggested the successful exclusion of chondroitin sulfate from peptides. Nitric oxide (NO) floods in cells activated by inflammation. It was inhibited when administrated with ESCH. To further explain the observed anti-inflammatory activity, ESCH was separated with Sephadex G-15 into 3 components, among which F3 showed a higher NO inhibition rate and significantly reduced the production of the proinflammatory cytokine IL-6. In addition, the yield of IL-10 increased. Western blotting suggested that F3 downregulated the NO content and IL-6 level by suppressing Mitogen-activated protein kinases (MAPK) channels. Moreover, both ESCH and F3 showed DPPH and ABTS free radical scavenging abilities which was possibly related to the anti-inflammatory property. These results indicated that ESCH behaved anti-inflammatory and antioxidant activities. Cartilage may be a good source to produce anti-inflammatory peptides.

GPER1 Modulates Synaptic Plasticity During the Development of Temporal Lobe Epilepsy in Rats.

G-protein coupled estrogen receptor 1 (GPER1) is a novel type of estrogen receptor. Several studies have shown that it has an anti-inflammatory action,which plays an important role in remyelination and cognitive ability adjustment. However, whether it is involved in the development of temporal lobe epilepsy (TLE) is still unknown. The present study established a TLE model by intraperitoneal injection of lithium chloride (3 mmol/kg) and pilocarpine (50 mg/kg) in rats to study the effect of GPER1 in the synaptic plasticity during the development of temporal lobe epilepsy. A microinjection cannula was implanted into the lateral ventricle region of rats via a stereotaxic instrument. G-1 is the specific GPER1 agonist and G15 is the specific GPER1 antagonist. The G1 or G15 and Dimethyl sulfoxide were injected into the rat brains in the intervention groups and control group, respectively. After G1 intervention, the learning and memory abilities and hippocampal neuron damage in epileptic rats were significantly improved, while G15 weakened the neuroprotective effect of GPER1. Meanwhile, G1 controlled the abnormal formation of hippocampal mossy fiber sprouting caused by seizures, and participated in the regulation of synaptic plasticity by reducing the expression of Synapsin I and increasing the expression of gephyrin. Inhibitory synapse gephyrin may play a significant role in synaptic plasticity.

The G-Protein-Coupled Estrogen Receptor (GPER) Regulates Trimethylation of Histone H3 at Lysine 4 and Represses Migration and Proliferation of Ovarian Cancer Cells In Vitro.

Histone H3 lysine 4 trimethylation (H3K4me3) is one of the most recognized epigenetic regulators of transcriptional activity representing, an epigenetic modification of Histone H3. Previous reports have suggested that the broad H3K4me3 domain can be considered as an epigenetic signature for tumor-suppressor genes in human cells. G-protein-coupled estrogen receptor (GPER), a new membrane-bound estrogen receptor, acts as an inhibitor on cell growth via epigenetic regulation in breast and ovarian cancer cells. This study was conducted to evaluate the relationship of GPER and H3K4me3 in ovarian cancer tissue samples as well as in two different cell lines (Caov3 and Caov4). Silencing of GPER by a specific siRNA and two selective regulators with agonistic (G1) and antagonistic (G15) activity were applied for consecutive in vitro studies to investigate their impacts on tumor cell growth and the changes in phosphorylated ERK1/2 (p-ERK1/2) and H3K4me3. We found a positive correlation between GPER and H3K4me3 expression in ovarian cancer patients. Patients overexpressing GPER as well as H3K4me3 had significantly improved overall survival. Increased H3K4me3 and p-ERK1/2 levels and attenuated cell proliferation and migration were observed in Caov3 and Caov4 cells via activation of GPER by G1. Conversely, antagonizing GPER activity by G15 resulted in opposite effects in the Caov4 cell line. In conclusion, interaction of GPER and H3K4me3 appears to be of prognostic significance for ovarian cancer patients. The results of the in vitro analyses confirm the biological rationale for their interplay and identify GPER agonists, such as G1, as a potential therapeutic approach for future investigations.

Molecular characterization of the first human G15 rotavirus strain of zoonotic origin from the bovine species.

Group A rotaviruses (RVAs) infect a wide variety of mammalian and avian species. Animals act as a potential reservoir to RVA human infections by direct virion transmission or by contributing genes to reassortants. Here, we report the molecular characterization of a rare human RVA strain Ni17-46 with a genotype G15P[14], isolated in Japan in 2017 during rotavirus surveillance in a paediatric outpatient clinic. The genome constellation of this strain was G15-P[14]-I2-R2-C2-M2-A13-N2-T9-E2-H3. This is the first report of an RVA with G15 genotype in humans, and sequencing and phylogenetic analysis results suggest that human infection with this strain has zoonotic origin from the bovine species. Given the fact that this strain was isolated from a patient with gastroenteritis and dehydration symptoms, we must take into account the virulence of this strain in humans.

Maximum expected entropy transformed Latin hypercube designs.

Existing projection designs (e.g. maximum projection designs) attempt to achieve good space-filling properties in all projections. However, when using a Gaussian process (GP), model-based design criteria such as the entropy criterion is more appropriate. We employ the entropy criterion averaged over a set of projections, called expected entropy criterion (EEC), to generate projection designs. We show that maximum EEC designs are invariant to monotonic transformations of the response, i.e. they are optimal for a wide class of stochastic process models. We also demonstrate that transformation of each column of a Latin hypercube design (LHD) based on a monotonic function can substantially improve the EEC. Two types of input transformations are considered: a quantile function of a symmetric Beta distribution chosen to optimize the EEC, and a nonparametric transformation corresponding to the quantile function of a symmetric density chosen to optimize the EEC. Numerical studies show that the proposed transformations of the LHD are efficient and effective for building robust maximum EEC designs. These designs give projections with markedly higher entropies and lower maximum prediction variances (MPV's) at the cost of small increases in average prediction variances (APV's) compared to state-of-the-art space-filling designs over wide ranges of covariance parameter values.

Phospholipase A2 group XV activity during cardiopulmonary bypass surgery.

OBJECTIVES: All patients who undergo cardiopulmonary bypass (CPB) experience some degree of ischemia reperfusion injury (IRI). Severe IRI-induced acute kidney injury (AKI) occurs in approximately 1-2% of patients undergoing CPB. Previous studies using activity-based protein profiling of urine identified group XV phospholipase A2, PLA2G15/LPLA2, as potentially associated with patients who develop AKI post CPB. The present study examined urinary PLA2G15/LPLA2 activity during CPB and in the near postoperative period for associations with subsequent AKI development. DESIGN & METHODS: Samples were collected in a nested case controlled cohort of 21 patients per group who either did (AKI) or did not (non-AKI) develop AKI post-operatively. Serum and urine samples from each patient before, during and after CPB were assayed for PLA2G15/LPLA2 activity. RESULTS: Urine activity significantly increased during the intra operative period. In contrast the activities in paired sera were markedly decreased during CPB. There was no correlation between the serum and urine activity levels of patients. There were no significant differences in activity levels of PLA2G15/LPLA2 in the urine or sera from patients that did and did not develop AKI. CONCLUSIONS: The lack of correlation between serum and urine activity levels suggests that the rapid intraoperative increases in PLA2G15/LPLA2 activity may originate from the kidney and as such offer an intraoperative indicator of early renal response to CPB associated stressors.

Long noncoding RNA RP11-757G1.5 sponges miR-139-5p and upregulates YAP1 thereby promoting the proliferation and liver, spleen metastasis of colorectal cancer.

BACKGROUND: Accumulating evidence indicates that long non-coding RNAs (lncRNAs) acting as crucial regulators in tumorigenesis. However, its biological functions of lncRNAs in colorectal cancer (CRC) have not been systematically clarified. METHODS: An unbiased screening was performed to identify disregulated lncRNAs revealed to be implicated in CRC carcinogenesis according to an online-available data dataset. In situ hybridization (ISH), RT-qPCR and RNA fluorescence in situ hybridization (RNA-FISH) were applied to detect RP11-757G1.5 expression in CRC tissues and cell lines. The associations of RP11-757G1.5 with clinicopathological characteristics were analyzed. Their effects on prognosis were analyzed by the Kaplan-Meier analysis, Log-rank test, Univariate and Multivariate Cox regression analysis. The potential biological function of RP11-757G1.5 in CRC was investigated by Colony formation, Edu cell proliferation, Flow cytometry, Wound healing and Transwell assays. Bioinformatics binding site analysis, Luciferase reporter assay, Ago2 immunoprecipitation assays, RNA pull-down assay, RT-qPCR and Western blotting were utilized to demonstrate the mechanism of RP11-757G1.5 acts as a molecular sponge of miR-139-5p to regulate the expression of YAP1. Finally, we further explore the potential role of RP11-757G1.5 in CRC orthotopic xenografts in vivo. RESULTS: We discovered a novel oncogenic lncRNA RP11-757G1.5, that was overexpressed in CRC tissues, especially in aggressive cases. Moreover, up-regulation of RP11-757G1.5 strongly correlated with poor clinical outcomes of patients with CRC. Functional analyses revealed that RP11-757G1.5 promoted cell proliferation in vitro and in vivo. Furthermore, RP11-757G1.5 stimulated cell migration and invasion in vitro and in vivo. Mechanistic studies illustrated that RP11-757G1.5 regulated the expression of YAP1 through sponging miR-139-5p and inhibiting its activity thereby promoting CRC progression and development. CONCLUSIONS: Altogether, these results reveal a novel RP11-757G1.5/miR-139-5p/YAP1 regulatory axis that participates in CRC carcinogenesis and progression.

Cisplatin resistance in gastric cancer cells is involved with GPR30-mediated epithelial-mesenchymal transition.

Cisplatin is the major chemotherapeutic drug in gastric cancer, particularly in treating advanced gastric cancer. Tumour cells often develop resistance to chemotherapeutic drugs, which seriously affects the efficacy of chemotherapy. GPR30 is a novel oestrogen receptor that is involved in the invasion, metastasis and drug resistance of many tumours. Targeting GPR30 has been shown to increase the drug sensitivity of breast cancer cells. However, few studies have investigated the role of GPR30 in gastric cancer. Epithelial-mesenchymal transition (EMT) has been shown to be associated with the development of chemotherapeutic drug resistance. In this study, we demonstrated that GPR30 is involved in cisplatin resistance by promoting EMT in gastric cancer. GPR30 knockdown resulted in increased sensitivity of different gastric cancer (GC) cells to cisplatin and alterations in the epithelial/mesenchymal markers. Furthermore, G15 significantly enhanced the cisplatin sensitivity of GC cells while G1 inhibited this phenomenon. In addition, EMT occurred when AGS and BGC-823 were treated with cisplatin. Down-regulation of GPR30 with G15 inhibited this transformation, while G1 promoted it. Taken together, these results revealed the role of GPR30 in the formation of cisplatin resistance, suggesting that targeting GPR30 signalling may be a potential strategy for improving the efficacy of chemotherapy in gastric cancer.

Neurologically Motivated Coupling Functions in Models of Motor Coordination.

We present an analysis of two Haken-Kelso-Bunz (HKB) oscillators coupled by a neurologically motivated function. We study the effect of time delay and weighted self-feedback and mutual feedback on the synchronization behavior of the model. We focus on identifying parameter regimes supporting experimentally observed decrease in oscillation amplitude and loss of anti-phase stability that has inspired the development of the HKB model. We show that a combination of cross-talk and nonlinearity in the coupling, along with physiologically relevant time delay, is able to quantitatively account for both drop in oscillation amplitude and loss of anti-phase stability in a frequency dependent manner. Furthermore, we demonstrate that the transition between discrete and rhythmic movements could be captured by this model. To this end, we carry out theoretical and numerical analysis of the emergence of in-phase and anti-phase oscillations.

Correcting an estimator of a multivariate monotone function with isotonic regression.

In many problems, a sensible estimator of a possibly multivariate monotone function may fail to be monotone. We study the correction of such an estimator obtained via projection onto the space of functions monotone over a finite grid in the domain. We demonstrate that this corrected estimator has no worse supremal estimation error than the initial estimator, and that analogously corrected confidence bands contain the true function whenever the initial bands do, at no loss to band width. Additionally, we demonstrate that the corrected estimator is asymptotically equivalent to the initial estimator if the initial estimator satisfies a stochastic equicontinuity condition and the true function is Lipschitz and strictly monotone. We provide simple sufficient conditions in the special case that the initial estimator is asymptotically linear, and illustrate the use of these results for estimation of a G-computed distribution function. Our stochastic equicontinuity condition is weaker than standard uniform stochastic equicontinuity, which has been required for alternative correction procedures. This allows us to apply our results to the bivariate correction of the local linear estimator of a conditional distribution function known to be monotone in its conditioning argument. Our experiments suggest that the projection step can yield significant practical improvements.