Prognostic Significance of Platelet (PLT) and Platelet to Mean Platelet Volume (PLT/MPV) Ratio During Apatinib Second-Line or Late-Line Treatment in Advanced Esophageal Squamous Cell Carcinoma Patients.

Background: Apatinib has a certain efficacy for advanced esophageal squamous cell carcinoma (ESCC). This study aimed to investigate the prognostic significance of platelet (PLT) and platelet to mean platelet volume (PLT/MPV) ratio for advanced ESCC patients with apatinib second-line or late-line treatment. Methods: A retrospective study included 80 patients with advanced ESCC who received Apatinib ≥ 2 lines targeted therapy. We collected baseline clinical characteristics and blood parameters from the patients. Kaplan-Meier plots and univariate and multivariate analysis were used to find the factors related to progression-free survival (PFS). Results: The optimal cut-off values of PLT and PLT/MPV ratio were determined by X-tile software. Kaplan-Meier analysis demonstrated that patients in the high PLT group had better PFS than those in the low PLT group (156 d vs 80 d, P <.001), and patients in the high PLT/MPV ratio group had better PFS than those in low PLT/MPV ratio group (157 d vs 85 d, P <.001). Univariate analysis revealed pretreatment PLT and PLT/MPV ratio were significantly correlated with PFS. Multivariate analysis revealed high levels of pretreatment PLT/MPV ratio was an independent predictor of longer PFS (HR: 0.257, 95% CI: 0.089-0.743, P = .012). Conclusion: High levels of baseline PLT and PLT/MPV may indicate a better prognosis in apatinib ≥ 2 lines treatment for advanced ESCC patients.

Combining Clinicopathology, IVIM-DWI and Texture Parameters for a Nomogram to Predict Treatment Response to Neoadjuvant Chemoradiotherapy in Locally Advanced Rectal Cancer Patients.

Objectives: This study aimed to create a nomogram for the risk prediction of neoadjuvant chemoradiotherapy (nCRT) resistance in locally advanced rectal cancer (LARC). Methods: Clinical data in this retrospective study were collected from a total of 135 LARC patients admitted to our hospital from June 2016 to December 2020. After screening by inclusion and exclusion criteria, 62 patients were included in the study. Texture analysis (TA) was performed on T2WI and DWI images. Patients were divided into response group (CR+PR) and no-response group (SD+PD) according to efficacy assessment. Multivariate analysis was performed on clinicopathology, IVIM-DWI and texture parameters for screening of independent predictors. A nomogram was created and model fit and clinical net benefit were assessed. Results: Multivariate analysis of clinicopathology parameters showed that the differentiation and T stage were independent predictors (OR values were 14.516 and 11.589, resp.; P<0.05). Multivariate analysis of IVIM-DWI and texture parameters showed that f value and Rads-score were independent predictors (OR values were 0.855, 2.790, resp.; P<0.05). In this study, clinicopathology together with IVIM-DWI and texture parameters showed the best predictive efficacy (AUC=0.979). The nomogram showed good predictive performance and stability in identifying high-risk LARC patients who are resistant to nCRT (C-index=0.979). Decision curve analyses showed that the nomogram had the best clinical net benefit. Ten-fold cross-validation results showed that the average AUC value was 0.967, and the average C-index was 0.966. Conclusions: The nomogram combining the differentiation, T stage, f value and Rads-score can effectively estimate the risk of nCRT resistance in patients with LARC.

In vitro free radical scavenging capacity of dimethylglycine sodium salt and its protective ability against oleic acid hydroperoxide-induced oxidative damage in IPEC-J2 cells.

The aim of the present study was to evaluate the in vitro free radical scavenging capacity of dimethylglycine sodium (DMG­Na) and its protective ability against oleic acid hydroperoxide (OAHPx)­induced oxidative damage in IPEC­J2 cells. Initially, the free radical scavenging activities of water­soluble pigments (DMG­Na, betalain, capsanthin and cyanidin­3­rutinoside) were measured and compared with those of Trolox. Subsequently, freshly collected swine blood was mixed with heparin and centrifuged to obtain erythrocytes. In order to induce the free radical chain oxidation in erythrocytes, the aqueous peroxyl radicals were generated by thermal decomposition of 2,2'­azobis(2­amidinopropane) dihydrochloride (AAPH) in oxygen. A 2% suspension of porcine erythrocytes in PBS buffer were pre­incubated for 30 min at 37˚C with DMG­Na (32 µM), followed by incubation with or without AAPH (75 mM) for 5 h with gentle shaking. Additionally, IPEC­J2 cells were randomly assigned to four groups (n=6 per group): Cells treated with phosphate buffered saline (PBS); cells treated with DMG­Na (32 µM); cells treated with oleic acid hydroperoxides (OAHPx, 20 µM; TO group); cells treated with DMG­Na (32 µM) followed by OAHPx (20 µM; DTO group). The cells were cultured in Dulbecco's modified Eagle's medium, Ham's F­12 mixture, 1.5 mM HEPES, 5% (v/v) fetal bovine serum, 1% (v/v) insulin­transferrin­selenium mixture, 1% (v/v) penicillin­streptomycin mixture and 2.5 µg/ml fungizone (37˚C, 5% CO2). The results showed that DMG­Na exerted the strongest free radical scavenging capacity at 0.32 M from 0.08­0.64 M, and that it could prevent AAPH­induced porcine erythrocyte hemolysis by increasing its antioxidant capacity (P<0.05). The results also demonstrated that antioxidant capacity and antioxidant­associated gene expression increased in the DTO group relative to the TO group (P<0.05), indicating that DMG­Na prevented the OAHPx­induced oxidative damage in IPEC­J2 cells by improving the antioxidant capacity and antioxidant­associated gene expression.

Characterization of retinal inosine monophosphate dehydrogenase 1 in several mammalian species.

PURPOSE: The purpose of this study was to characterize the inosine monophosphate dehydrogenase 1 (IMPDH1) protein isoforms in mammalian retinas, in order to determine the species distribution of these variants and identify an optimal animal model for studying IMPDH1-associated retinal diseases. Mutations in IMPDH1 cause the RP10 form of autosomal dominant retinitis pigmentosa, and are a rare cause of Leber congenital amaurosis. METHODS: Retinas from several mammalian species were obtained commercially. Human retinas were isolated by the San Diego Eye Bank and flash frozen within four hours post mortem. Proteins were isolated from retinal tissue using the PARIS protocol. Anti-IMPDH1 antibodies were used to visualize the IMDPH1 proteins on Western blots. RESULTS: Transcript and protein analyses have shown that IMPDH1 undergoes alternate splicing to produce at least two retinal isoforms in both human and mouse. The relative abundance of these IMPDH1 isoforms is different between mouse and human. This study extends these findings by showing that the two IMPDH1 isoforms are also present in dog, rat, sheep, pig, and cow retina, but that, as with mouse, the relative abundances of these isoforms differ from those found in human retina. CONCLUSIONS: The existence of two major retinal isoforms of the IMPDH1 protein is maintained across all mammalian species tested. The relative abundance of IMPDH1 proteins in human retina is unique in comparison to other mammalian species, indicating an apparent lack of an ideal model organism for human retinal IMPDH1 expression. Pig and/or sheep may prove to be potential model organisms based on the observed retinal isoform abundance in these species. These findings will aid future research in understanding the role of retinal-specific IMPDH1 proteins, and will contribute to research elucidating the pathophysiology associated with IMPDH1 missense mutations.

Why do mutations in the ubiquitously expressed housekeeping gene IMPDH1 cause retina-specific photoreceptor degeneration?

PURPOSE: The purpose of this study was to investigate retinal inosine monophosphate dehydrogenase 1 (IMPDH1) transcripts and proteins to gain an understanding of how mutations in IMPDH1 lead to retinal disease. Mutations in IMPDH1 cause the RP10 form of autosomal dominant retinitis pigmentosa (adRP) and are a rare cause of dominant Leber congenital amaurosis (LCA). IMPDH1 is a highly conserved, widely expressed housekeeping gene, the product of which catalyzes the rate-limiting step of de novo guanine synthesis. Despite its conservation and ubiquity, the clinical consequences of missense mutations in IMPDH1 are limited to the retina, and the disease mechanism is currently unknown. METHODS: A variety of methods were used to address the unique features of IMPDH1 in the retina, including Northern blot analysis, serial analysis of gene expression (SAGE), immunohistochemistry, transcript sequencing, and Western blot analysis. RESULTS: Results of the experiments showed that IMPDH1 levels are higher in the retina than in any other tissue tested. Specifically, IMPDH1 is found predominately in the inner segment and synaptic terminals of retinal photoreceptors. The predominant transcripts of IMPDH1 in human retina are the result of alternate splicing and alternate start sites of translation. They are significantly different from those in other tissues, and these variant transcripts encode distinct proteins. Further, the proportions of IMPDH1 transcripts and proteins in human retina are different from those in mouse retina. CONCLUSIONS: Identification of unique retinal isoforms supports the existence of a novel IMPDH1 function in the retina, one that is probably altered by disease-causing mutations. This alone, or coupled with the high levels of IMPDH1 in the retina, may explain the retina-specific phenotype associated with IMPDH1 mutations. Elucidating the functional properties of these unique, human retinal isoforms is crucial to understanding the pathophysiology of IMPDH1 mutations.

Spectrum and frequency of mutations in IMPDH1 associated with autosomal dominant retinitis pigmentosa and leber congenital amaurosis.

PURPOSE: The purpose of this study was to determine the frequency and spectrum of inosine monophosphate dehydrogenase type I (IMPDH1) mutations associated with autosomal dominant retinitis pigmentosa (RP), to determine whether mutations in IMPDH1 cause other forms of inherited retinal degeneration, and to analyze IMPDH1 mutations for alterations in enzyme activity and nucleic acid binding. METHODS: The coding sequence and flanking intron/exon junctions of IMPDH1 were analyzed in 203 patients with autosomal dominant RP (adRP), 55 patients with autosomal recessive RP (arRP), 7 patients with isolated RP, 17 patients with macular degeneration (MD), and 24 patients with Leber congenital amaurosis (LCA). DNA samples were tested for mutations by sequencing only or by a combination of single-stranded conformational analysis and by sequencing. Production of fluorescent reduced nicotinamide adenine dinucleotide (NADH) was used to measure enzymatic activity of mutant IMPDH1 proteins. The affinity and the specificity of mutant IMPDH1 proteins for single-stranded nucleic acids were determined by filter-binding assays. RESULTS: Five different IMPDH1 variants, Thr116Met, Asp226Asn, Val268Ile, Gly324Asp, and His 372Pro, were identified in eight autosomal dominant RP families. Two additional IMPDH1 variants, Arg105Trp and Asn198Lys, were found in two patients with isolated LCA. None of the novel IMPDH1 mutants identified in this study altered the enzymatic activity of the corresponding proteins. In contrast, the affinity and/or the specificity of single-stranded nucleic acid binding were altered for each IMPDH1 mutant except the Gly324Asp variant. CONCLUSIONS: Mutations in IMPDH1 account for approximately 2% of families with adRP, and de novo IMPDH1 mutations are also rare causes of isolated LCA. This analysis of the novel IMPDH1 mutants substantiates previous reports that IMPDH1 mutations do not alter enzyme activity and demonstrates that these mutants alter the recently identified single-stranded nucleic acid binding property of IMPDH. Studies are needed to further characterize the functional significance of IMPDH1 nucleic acid binding and its potential relationship to retinal degeneration.

Prevalence of disease-causing mutations in families with autosomal dominant retinitis pigmentosa: a screen of known genes in 200 families.

PURPOSE: To survey families with clinical evidence of autosomal dominant retinitis pigmentosa (adRP) for mutations in genes known to cause adRP. METHODS: Two hundred adRP families, drawn from a cohort of more than 400 potential families, were selected by analysis of pedigrees. Minimum criteria for inclusion in the adRP cohort included either evidence of at least three generations of affected individuals or two generations with evidence of male-to-male transmission. Probands from each family were screened for mutations in 13 genes known to cause adRP: CA4, CRX, FSCN2, IMPDH1, NRL, PRPF3 (RP18), PRPF8 (RP13), PRPF31 (RP11), RDS, RHO, ROM1, RP1, and RP9. Families without mutations in autosomal genes and in which an X-linked mode of inheritance could not be excluded were tested for mutations in ORF 15 of X-linked RPGR. Potentially pathogenic variants were evaluated based on a variety of genetic and computational criteria, to confirm or exclude pathogenicity. RESULTS: A total of 82 distinct, rare (nonpolymorphic) variants were detected among the genes tested. Of these, 57 are clearly pathogenic based on multiple criteria, 10 are probably pathogenic, and 15 are probably benign. In the cohort of 200 families, 94 (47%) have one of the clearly pathogenic variants and 10 (5%) have one of the probably pathogenic variants. One family (0.5%) has digenic RDS-ROM1 mutations. Two families (1%) have a pathogenic RPGR mutation, indicating that families with apparent autosomal transmission of RP may actually have X-linked genetic disease. Thus, 107 families (53.5%) have mutations in known genes, leaving 93 whose underlying cause is still unknown. CONCLUSIONS: Together, the known adRP genes account for retinal disease in approximately half of the families in this survey, mostly Americans of European origin. Among the adRP genes, IMPDH1, PRPF8, PRPF31, RDS, RHO, and RP1 each accounts for more than 2% of the total; CRX, PRPF3, and RPGR each accounts for roughly 1%. Disease-causing mutations were not found in CA4, FSCN2, NRL, or RP9. Because some mutations are frequent and some regions are more likely to harbor mutations than others, more than two thirds of the detected mutations can be found by screening less than 10% of the total gene sequences. Among the remaining families, mutations may lie in regions of known genes that were not tested, mutations may not be detectable by PCR-based sequencing, or other loci may be involved.

Adenovirus or HA-2 fusogenic peptide-assisted lipofection increases cytoplasmic levels of plasmid in nondividing endothelium with little enhancement of transgene expression.

BACKGROUND: Adenovirus-assisted lipofection has been reported to increase transfection efficiency through mechanisms potentially involving endosome escape and/or nuclear targeting activity. Similarly, transfection with the viral fusogenic peptide HA-2 of the influenza virus hemagglutinin can increase transfection efficiency. However, there are few studies examining the mechanism and intracellular trafficking of these viral and/or viral fusogenic peptide-assisted lipofections. METHODS AND RESULTS: Endosome escape was directly assayed with T7 RNA polymerase bound to plasmid (pTM beta gal) expressing beta-galactosidase under a T7 promoter to detect transcribable plasmid that escapes the endosomal compartment. Lipofection of pTM beta gal with replication-deficient adenovirus (Ad5-null) at a multiplicity of infection (MOI) of 100 and 1000 increased cytoplasmic levels of transcribable plasmid by 24- and 117-fold, respectively, over lipofection alone, without an effect on total plasmid uptake. However, lipofection of pCMV beta gal with Ad5-null at a MOI of 100 and 1000 increased transgene expression only seven- and eight-fold, respectively, over lipofection alone. Thus, a 24-fold increase in endosome escape saturated expression from pCMV beta gal and provided only a seven-fold benefit in nondividing cells, which was not significantly increased with further increases in endosome escape. A cationic form of HA-2 (HA-K(4)) also caused significant enhancements in endosome escape, as detected with the cytoplasmic transcription assay. However, HA-K(4) enhancement of endosome escape did not correlate with transgene expression from pCMV beta gal, consistent with the detection of HA-K(4)-mediated partitioning of plasmid to the insoluble fraction of the cell lysate. CONCLUSION: These results indicate that enhancement of endosome escape in nondividing cells does not fully alleviate rate limits related to nuclear import of the plasmid.