The role of adenocarcinoma subtypes and immunohistochemistry in predicting lymph node metastasis in early invasive lung adenocarcinoma.

BACKGROUND: Identifying lymph node metastasis areas during surgery for early invasive lung adenocarcinoma remains challenging. The aim of this study was to develop a nomogram mathematical model before the end of surgery for predicting lymph node metastasis in patients with early invasive lung adenocarcinoma. METHODS: In this study, we included patients with invasive lung adenocarcinoma measuring ≤ 2 cm who underwent pulmonary resection with definite pathology at Qilu Hospital of Shandong University from January 2020 to January 2022. Preoperative biomarker results, clinical features, and computed tomography characteristics were collected. The enrolled patients were randomized into a training cohort and a validation cohort in a 7:3 ratio. The training cohort was used to construct the predictive model, while the validation cohort was used to test the model independently. Univariate and multivariate logistic regression analyses were performed to identify independent risk factors. The prediction model and nomogram were established based on the independent risk factors. Recipient operating characteristic (ROC) curves were used to assess the discrimination ability of the model. Calibration capability was assessed using the Hosmer-Lemeshow test and calibration curves. The clinical utility of the nomogram was assessed using decision curve analysis (DCA). RESULTS: The overall incidence of lymph node metastasis was 13.23% (61/461). Six indicators were finally determined to be independently associated with lymph node metastasis. These six indicators were: age (P < 0.001), serum amyloid (SA) (P = 0.008); carcinoma antigen 125 (CA125) (P = 0. 042); mucus composition (P = 0.003); novel aspartic proteinase of the pepsin family A (Napsin A) (P = 0.007); and cytokeratin 5/6 (CK5/6) (P = 0.042). The area under the ROC curve (AUC) was 0.843 (95% CI: 0.779-0.908) in the training cohort and 0.838 (95% CI: 0.748-0.927) in the validation cohort. the P-value of the Hosmer-Lemeshow test was 0.0613 in the training cohort and 0.8628 in the validation cohort. the bias of the training cohort corrected C-index was 0.8444 and the bias-corrected C-index for the validation cohort was 0.8375. demonstrating that the prediction model has good discriminative power and good calibration. CONCLUSIONS: The column line graphs created showed excellent discrimination and calibration to predict lymph node status in patients with ≤ 2 cm invasive lung adenocarcinoma. In addition, the predictive model has predictive potential before the end of surgery and can inform clinical decision making.

Prognostic significance and survival benefits of postoperative adjuvant chemotherapy in patients with stage IA lung adenocarcinoma with non-predominant micropapillary components.

BACKGROUND: The prognostic significance of adjuvant chemotherapy (ACT) for patients with stage IA micropapillary non-predominant (MPNP) lung adenocarcinoma (LUAD) remains unknown. This study aimed to investigate the effects of postoperative ACT in patients with stage IA MPNP-LUAD. METHODS: A total of 149 patients with pathological stage IA MPNP-LUAD who underwent surgery at our center were retrospectively analyzed. Propensity score matching (PSM) analysis was conducted to reduce potential selection bias. Kaplan-Meier analyses were used to assess the impact of ACT on recurrence-free survival (RFS), overall survival (OS), and disease-specific survival (DSS). Subgroup analyses were performed for the survival outcomes based on the percentage of micropapillary components. Cox proportional hazards regression analyses were applied to identify risk factors associated with survival. RESULTS: The receipt or non-receipt of postoperative ACT had no significant effect on RFS, OS, and DSS among all enrolled patients with stage IA MPNP-LUAD (P > 0.05). For patients with a micropapillary component > 5%, the 5-year rates of RFS, OS, and DSS were significantly higher in the ACT group compared to the observation group, both before and after PSM (P < 0.05). However, the differences between the two groups were not significant for patients with a micropapillary component ≤ 5% (P > 0.05). The resection range (HR = 0.071; 95% CI: 0.020-0.251; P < 0.001), tumor size (HR = 2.929; 95% CI: 1.171-7.330; P = 0.022), and ACT (HR = 0.122; 95% CI: 0.037-0.403; P = 0.001) were identified as independent prognostic factors for RFS through Cox regression analysis. CONCLUSION: Patients with stage IA MPNP-LUAD who have a micropapillary component greater than 5% might benefit from postoperative ACT, while those with a micropapillary component ≤ 5% did not appear to derive the same benefit from postoperative ACT.

Construction and validation of a predictive model of invasive adenocarcinoma in pure ground-glass nodules less than 2 cm in diameter.

OBJECTIVES: In this study, we aimed to develop a multiparameter prediction model to improve the diagnostic accuracy of invasive adenocarcinoma in pulmonary pure glass nodules. METHOD: We included patients with pulmonary pure glass nodules who underwent lung resection and had a clear pathology between January 2020 and January 2022 at the Qilu Hospital of Shandong University. We collected data on the clinical characteristics of the patients as well as their preoperative biomarker results and computed tomography features. Thereafter, we performed univariate and multivariate logistic regression analyses to identify independent risk factors, which were then used to develop a prediction model and nomogram. We then evaluated the recognition ability of the model via receiver operating characteristic (ROC) curve analysis and assessed its calibration ability using the Hosmer-Lemeshow test and calibration curves. Further, to assess the clinical utility of the nomogram, we performed decision curve analysis. RESULT: We included 563 patients, comprising 174 and 389 cases of invasive and non-invasive adenocarcinoma, respectively, and identified seven independent risk factors, namely, maximum tumor diameter, age, serum amyloid level, pleural effusion sign, bronchial sign, tumor location, and lobulation. The area under the ROC curve was 0.839 (95% CI: 0.798-0.879) for the training cohort and 0.782 (95% CI: 0.706-0.858) for the validation cohort, indicating a relatively high predictive accuracy for the nomogram. Calibration curves for the prediction model also showed good calibration for both cohorts, and decision curve analysis showed that the clinical prediction model has clinical utility. CONCLUSION: The novel nomogram thus constructed for identifying invasive adenocarcinoma in patients with isolated pulmonary pure glass nodules exhibited excellent discriminatory power, calibration capacity, and clinical utility.

[Characteristics of Inflammatory Markers in Diabetic Foot Patients and Their Relationship With Prognosis of Diabetic Foot Ulcers]. / ç³–å°¿ç— è¶³æ‚£è€ ç‚Žç—‡æ ‡å¿—ç‰©ç‰¹å¾åŠå ¶ä¸Žè¶³æºƒç–¡é¢„åŽå ³ç³».

Objective: To explore the characteristics of baseline inflammatory markers in diabetic foot patients and their relationship with the prognosis of diabetic foot ulcers. Methods: The clinical data of diabetic foot patients (n=495) admitted to West China Hospital, Sichuan University since 2016 were retrospectively collected through the hospital electronic medical record system to analyze the characteristics of inflammatory markers and their relationship with the prognosis of diabetic foot ulcers. Results: White blood cell count (WBC), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), procalcitonin (PCT), and interleukin-6 (IL-6) levels were significantly higher in patients defined as grade 4 on the Wagner Scale than those in patients defined as grade 0-3 on the Wagner Scale. Neutrophil percentage (NE%) was higher in Wagner grade-4 patients than those in Wagner grade-0 and grade-1 patients and higher in Wagner grade-3 patients than those in Wagner grade-0 patients. NE%, CRP, PCT, and IL-6 levels were positively correlated with the severity of diabetic foot, with the respective odds ratio (OR) at 95% confidence interval (CI) being 1.038 (1.019-1.056), 1.019 (1.012-1.026), 8.225 (2.015-33.576), and 1.017 (1.008-1.025). Using Wagner grade-0 patients as the reference, patients with higher WBC were more likely to progress to Wagner grade 2, 3, and 4, with the respective OR (95% CI) values being 1.260 (1.096-1.447), 1.188 (1.041-1.356), and 1.301 (1.137-1.490); patients with higher ESR were more likely to progress to Wagner grade 3 and 4, with the respective OR (95% CI) values being 1.030 (1.006-1.054) and 1.045 (1.019-1.071). Baseline ESR (P=0.008), CRP (P=0.039), and IL-6 (P=0.033) levels were lower in patients who had received antibiotics prior to their admission than those in patients who had not received antibiotics before admission. The levels of WBC, NE%, ESR, PCT, and IL-6 were lower in the full recovery group than those in the group of patients who did not respond to treatment. The higher the levels of NE% and IL-6, the worse the prognosis of diabetic foot ulcers became, with the respective OR (95% CI) values being 1.030 (1.010-1.051) and 1.008 (1.002-1.013). Conclusion: The severity of diabetic foot ulcers increased with the rise in baseline levels of inflammatory markers. Elevated baseline NE% and IL-6 levels suggest a poor prognosis. Our findings suggest that early assessment of diabetic foot infection and standardized antibiotic therapy should be implemented to improve the prognosis.

First report of alfalfa root rot caused by Trichothecium roseum in China.

Alfalfa (Medicago sativa L.) is perennial leguminous forage, which is cultivated throughout the world due to its high yield, high quality, satisfactory palatability, and wide adaptability. With the increase of planting area in China, root diseases caused by Fusarium spp., Sclerotium rolfsii, Phytophthora spp. (Yang et al. 2022), and new pathogens have been found that reduce the yield and quality of alfalfa and cause economic losses (Li at al. 2019). In 2021, an alfalfa disease occurred under conditions of high temperature and high humidity at the Jiaozhou Experimental Base of Qingdao Agricultural University (Jiaozhou Modern Agricultural Science and Technology Demonstration Park, 36.33°N 120.40°E, Qingdao, Shandong, China), and about 2 ha of alfalfa were infected. The disease affected up to 35% of the plants and caused grass spots. Infected plants developed black-brown lesions with irregular shapes on roots with yellowing of the foliage; the leaves of the whole plant turned yellow. In the late stage of the disease, defoliation occurred and the plants stopped growing, wilted and died. Ten infected plants with typical symptom were collected for isolation and identification of pathogen. The infected roots were cut into 3-5 mm2 sections and then soaked in 75% ethanol for 30 s, followed by a 3-minute immersion in 2% sodium hypochlorite for surface sterilization. Next, the tissues were rinsed in sterile water five times and then placed on potato dextrose agar (PDA) medium. After three subcultures and subsequent single spore isolation, one representative strain named as DC1 was isolated from the infected roots. Based on morphological observation, the colony of DC1 was flat, granular, and powdery in appearance. Four days after inoculation on PDA medium, the size of the colony were 2.1-2.6 cm. After 8 to 20 days, the colonies were initially white and gradually change a light pink to peach color. The conidia are two-celled (Hamid et al. 2014), elliptic to pear-shaped, colorless or translucent, smooth to slightly rough with thick walls. The size of conidia ranged from 11.3 to 23.5 µm long × 6.1 to 12.7 µm wide (n =30). For the identification, the rDNA--ITS gene of the fungus was amplified using the primers ITS1/ITS4 (White et al.1990), and the EF1α gene was amplified using primers EF1-983F/EF1-2218R (Rehner and Buckley 2005). Then the PCR amplicons were cloned into the pCE2 TA/Blunt-Zero vector. The results of the rDNA-ITS (OM049197.1, 515 bp) and EF1α (OM069381.1, 926 bp) sequences were deposited in GenBank. DNA analysis showed that the two sequences were 100% similar to the rDNA-ITS sequence (MN882763.1) and EF1α sequence (DQ676610.1) of Trichothecium roseum, respectively. A pathogenicity test was done by placing one piece (0.5 cm in diameter) of fungal culture (PDA plug) 1cm below the crown of 40-day-old healthy alfalfa (cv. Zhongmu No.3) plants, 3 replicates and 20 plants in each replicate. PDA plug without the pathogen were used for control. All plants were cultivated in a growth chamber at 25±1°C with a light cycle of 15 h (90% relative humidity). After 18 days, the roots of inoculated plants had dark brown lesions and the leaves of their plants turn yellow, while those control plants had no symptoms. To fulfilling Koch's postulates, the same pathogen was re-isolated from necrotic root tissue of inoculated plants and confirmed by morphology and the rDNA-ITS and EF1-α sequences. Based on disease symptoms, morphological characteristics DNA sequences and pathogenicity, the pathogen of alfalfa disease in Jiaozhou Experimental Base of Qingdao Agricultural University was identified as T. roseum. To our knowledge, this is first report of T. roseum causing alfalfa root rot. The newly emerging disease may pose threat to alfalfa production of central and southern China in future.

Southern Blight on Medicago sativa Caused by Sclerotium rolfsii in North China.

Alfalfa (Medicago sativa L.) is one of the most important perennial leguminous forages in many countries, known by its high feed value and yield potential. With the increasing demand for feed, alfalfa has been planted all over China. However, an increasingly serious alfalfa disease was observed and may restrict the development of the alfalfa industry in North China. In August 2019, an emerging alfalfa disease with symptoms resembling southern blight was observed in Jiaozhou experimental base (Jiaozhou Modern Agricultural Science and Technology Demonstration Park) of Qingdao Agricultural University (Qindao, Shandong province, China). The infected plants showed dark brown lesions on the stems and yellowing and wilting of the leaves. The pathogen produced white fluffy mycelia, and later sclerotia on stems and roots; the disease affected up to 25% of the plants and causes bare spots filled with weeds (Figure S1). Typical symptomatic tissues were brought back to the laboratory for pathogen isolation and identification. Fragments (3-5mm2) of root tissues were excised from lesions on the symptomatic roots and their surfaces were disinfested by sequential dipping in 70% ethanol for 30 s and in 2% NaClO for 3 min, then the fragments were rinsed in sterile water five times and cultured on potato dextrose (PDA) medium amended with streptomycin sulfate (0.1mg/mL). Cultures were incubated at 28°C in the dark and purified in PDA medium for three times. A representative strain (coded as CZL1) was isolated from the root rot of the diseased plant. After four days incubation on PDA, CZL1 formed white fluffy aerial mycelium 5.6-6 cm in diameter typical of S. rolfsii. After 15 to 20 days, abundant round sclerotia approximately1-3 mm in diameter were produced on the surface of the culture (Figure S2). The sclerotia were white at first and then gradually turned dark brown. To confirm the identity of the causal fungus, the complete internal transcribed spacer (ITS) rDNA region of the fungus was amplified using the primers ITS1/ITS4 (White et al.1990), and the elongation factor-1a gene (EF1a) was amplified using primers EF1-983F/EF1-2218R (Rehner and Buckley 2005). Then the PCR amplicons were cloned into the pCE2 TA/Blunt-Zero vector. The isolate was determined to contain two distinct sequence types for each gene. The results of ITS (MT812692, MT812693) and EF1a (MT846496 and MT846497) sequences were deposited in GenBank. DNA analysis revealed that the two ITS sequences were more than 99% identical to Athelia rolfsii (MN872304) in the NCBI GenBank database, and two EF1a sequences were 99% identical to the A. rolfsii EF1a sequence MN702789 and KP982854. To fulfill Koch's postulates, infected sorghum grain was placed near the roots of 15 40-day-old healthy alfalfa seedlings split into 3 pots with the same number of seedlings receiving a control treatment of sterilized sorghum grain. All plants were incubated in growth chamber at 24±1°C with 14-h-photoperiod (85% relative humidity). After 10-15 days, blight symptoms identical to those in the field were observed on inoculated plants, whereas those control plants were symptomless (Figure S2). S. rolfsii was successfully re-isolated from the inoculated plants and molecularly characterized as described above. Based on disease symptoms, fungal colonies, the ITS and EF1a sequence, and pathogenicity to the host, this fungus was identified as S. rolfsii (teleomorph Athelia rolfsii). To our knowledge, this is the first report of S. rolfsii as the causal agent of southern blight of alfalfa in North China, and it is also the first report of southern blight on alfalfa caused by S. rolfsii in China since 1996 observed in Guizhou province (Mo and Luo 1996).

Comprehensive analysis of coding sequence architecture features and gene expression in Arachis duranensis.

Coding sequence (CDS) architecture affects gene expression levels in organisms. Codon optimization can increase the gene expression level. Therefore, understanding codon usage patterns has important implications for research on genetic engineering and exogenous gene expression. To date, the codon usage patterns of many model plants have been analyzed. However, the relationship between CDS architecture and gene expression in Arachis duranensis remains poorly understood. According to the results of genome sequencing, A. duranensis has many resistant genes that can be used to improve the cultivated peanut. In this study, bioinformatic approaches were used to estimate A. duranensis CDS architectures, including frequency of the optimal codon (Fop), polypeptide length and GC contents at the first (GC1), second (GC2) and third (GC3) codon positions. In addition, Arachis RNA-seq datasets were downloaded from PeanutBase. The relationships between gene expression and CDS architecture were assessed both under normal growth as well as nematode and drought stress conditions. A total of 26 codons with high frequency were identified, which preferentially ended with A or T in A. duranensis CDSs under the above-mentioned three conditions. A similar CDS architecture was found in differentially expressed genes (DEGs) under nematode and drought stresses. The GC1 content differed between DEGs and non-differentially expressed genes (NDEGs) under both drought and nematode stresses. The expression levels of DEGs were affected by different CDS architectures compared with NDEGs under drought stress. In addition, no correlation was found between differential gene expression and CDS architecture neither under nematode nor under drought stress. These results aid the understanding of gene expression in A. duranensis.

Metabolomic analyses reveal substances that contribute to the increased freezing tolerance of alfalfa (Medicago sativa L.) after continuous water deficit.

BACKGROUND: Alfalfa is a high-quality forage cultivated widely in northern China. Recently, the failure of alfalfa plants to survive the winter has caused substantial economic losses. Water management has attracted considerable attention as a method for the potential improvement of winter survival. The aim of this study was to determine whether and how changes in the water regime affect the freezing tolerance of alfalfa. RESULTS: The alfalfa variety WL353LH was cultivated under water regimes of 80 and 25% of water-holding capacity, and all the plants were subjected to low temperatures at 4/0 °C (light/dark) and then - 2/- 6 °C (light/dark). The semi-lethal temperatures were lower for water-stressed than well-watered alfalfa. The pool sizes of total soluble sugars, total amino acids, and proline changed substantially under water-deficit and low-temperature conditions. Metabolomics analyses revealed 72 subclasses of differential metabolites, among which lipid and lipid-like molecules (e.g., fatty acids, unsaturated fatty acids, and glycerophospholipids) and amino acids, peptides, and analogues (e.g., proline betaine) were upregulated under water-deficit conditions. Some carbohydrates (e.g., D-maltose and raffinose) and flavonoids were also upregulated at low temperatures. Finally, Kyoto Encyclopedia of Genes and Genomes analyses revealed 18 significantly enriched pathways involved in the biosynthesis and metabolism of carbohydrates, unsaturated fatty acids, amino acids, and glycerophospholipids. CONCLUSIONS: Water deficit significantly enhanced the alfalfa' freezing tolerance, and this was correlated with increased soluble sugar, amino acid, and lipid and lipid-like molecule contents. These substances are involved in osmotic regulation, cryoprotection, and the synthesis, fluidity, and stability of the cellular membrane. Our study provides a reference for improving alfalfa' winter survival through water management.

Analysis of physiological and miRNA responses to Pi deficiency in alfalfa (Medicago sativa L.).

KEY MESSAGE: The induction of miR399 and miR398 and the inhibition of miR156, miR159, miR160, miR171, miR2111, and miR2643 were observed under Pi deficiency in alfalfa. The miRNA-mediated genes involved in basic metabolic process, root and shoot development, stress response and Pi uptake. Inorganic phosphate (Pi) deficiency is known to be a limiting factor in plant development and growth. However, the underlying miRNAs associated with the Pi deficiency-responsive mechanism in alfalfa are unclear. To elucidate the molecular mechanism at the miRNA level, we constructed four small RNA (sRNA) libraries from the roots and shoots of alfalfa grown under normal or Pi-deficient conditions. In the present study, alfalfa plants showed reductions in biomass, photosynthesis, and Pi content and increases in their root-to-shoot ratio and citric, malic, and succinic acid contents under Pi limitation. Sequencing results identified 47 and 44 differentially expressed miRNAs in the roots and shoots, respectively. Furthermore, 909 potential target genes were predicted, and some targets were validated by RLM-RACE assays. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed prominent enrichment in signal transducer activity, binding and basic metabolic pathways for carbohydrates, fatty acids and amino acids; cellular response to hormone stimulus and response to auxin pathways were also enriched. qPCR results verified that the differentially expressed miRNA profile was consistent with sequencing results, and putative target genes exhibited opposite expression patterns. In this study, the miRNAs associated with the response to Pi limitation in alfalfa were identified. In addition, there was an enrichment of miRNA-targeted genes involved in biological regulatory processes such as basic metabolic pathways, root and shoot development, stress response, Pi transportation and citric acid secretion.

Molecular cloning and functional analysis of the drought tolerance gene MsHSP70 from alfalfa (Medicago sativa L.).

Heat shock proteins (HSPs) are a ubiquitously expressed class of protective proteins that play a key role in plant response to stressful conditions. This study aimed to characterize and investigate the function of an HSP gene in alfalfa (Medicago sativa). MsHSP70, which contains a 2028-bp open reading frame, was identified through homology cloning. MsHSP70 shares high sequence identity (94.47%) with HSP70 from Medicago truncatula. Expression analysis of MsHSP70 in alfalfa organs revealed a relatively higher expression level in aerial organs such as flowers, stems and leaves than in roots. MsHSP70 was induced by heat shock, abscisic acid (ABA) and hydrogen peroxide. Transgenic Arabidopsis seedlings overexpressing MsHSP70 were hyposensitive to polyethylene glycol (PEG) and ABA treatments, suggesting that exogenous expression of MsHSP70 enhanced Arabidopsis tolerance to these stresses. Examination of physiological indexes related to drought and ABA stress demonstrated that in comparison with non-transgenic plants, T3 transgenic Arabidopsis plants had an increased proline content, higher superoxide dismutase (SOD) activity, and decreased malondialdehyde (MDA) content. Furthermore, higher relative water content (RWC) was detected in transgenic plants compared with non-transgenic plants under drought stress. These findings clearly indicate that molecular manipulation of MsHSP70 in plants can have substantial effects on stress tolerance.

Silver-Catalyzed Cascade Radical Cyclization: A Direct Approach to 3,4-Disubstituted Dihydroquinolin-2(1H)-ones through Activation of the P-H Bond and Functionalization of the C(sp(2))-H Bond.

A silver-catalyzed cascade cyclization of cinnamamides with diphenylphosphine oxide was developed, in which activation of the P-H bond and functionalization of the C(sp(2))-H bond occurred. A direct method for the synthesis of 3,4-disubstituted dihydroquinolin-2(1H)-ones was developed.

Molecular cloning and characterization of the MsHSP17.7 gene from Medicago sativa L.

Heat shock proteins (HSPs) are ubiquitous protective proteins that play crucial roles in plant development and adaptation to stress, and the aim of this study is to characterize the HSP gene in alfalfa. Here we isolated a small heat shock protein gene (MsHSP17.7) from alfalfa by homology-based cloning. MsHSP17.7 contains a 477-bp open reading frame and encodes a protein of 17.70-kDa. The amino acid sequence shares high identity with MtHSP (93.98 %), PsHSP17.1 (83.13 %), GmHSP17.9 (74.10 %) and SlHSP17.6 (79.25 %). Phylogenetic analysis revealed that MsHSP17.7 belongs to the group of cytosolic class II small heat shock proteins (sHSP), and likely localizes to the cytoplasm. Quantitative RT-PCR indicated that MsHSP17.7 was induced by heat shock, high salinity, peroxide and drought stress. Prokaryotic expression indicated that the salt and peroxide tolerance of Escherichia coli was remarkably enhanced. Transgenic Arabidopsis plants overexpressing MsHSP17.7 exhibited increased root length of transgenic Arabidopsis lines under salt stress compared to the wild-type line. The malondialdehyde (MDA) levels in the transgenic lines were significantly lower than in wild-type, although proline levels were similar between transgenic and wild-type lines. MsHSP17.7 was induced by heat shock, high salinity, oxidative stress and drought stress. Overexpression analysis suggests that MsHSP17.7 might play a key role in response to high salinity stress.

Isolation and functional characterization of salt-stress induced RCI2-like genes from Medicago sativa and Medicago truncatula.

Salt stress is one of the most significant adverse abiotic factors, causing crop failure worldwide. So far, a number of salt stress-induced genes, and genes improving salt tolerance have been characterized in a range of plants. Here, we report the isolation and characterization of a salt stress-induced Medicago sativa (alfalfa) gene (MsRCI2A), which showed a high similarity to the yeast plasma membrane protein 3 gene (PMP3) and Arabidopsis RCI2A. The sequence comparisons revealed that five genes of MtRCI2(A-E) showed a high similarity to MsRCI2A in the Medicago truncatula genome. MsRCI2A and MtRCI2(A-E) encode small, highly hydrophobic proteins containing two putative transmembrane domains, predominantly localized in the plasma membrane. The transcript analysis results suggest that MsRCI2A and MtRCI2(A-D) genes are highly induced by salt stress. The expression of MsRCI2A and MtRCI2(A-C) in yeast mutants lacking the PMP3 gene can functionally complement the salt sensitivity phenotype resulting from PMP3 deletion. Overexpression of MsRCI2A in Arabidopsis plants showed improved salt tolerance suggesting the important role of MsRCI2A in salt stress tolerance in alfalfa.

Fabrication, structure, characterization and emulsion application of citrate agar.

In this study, citric acid successfully reacted with agar through the dry heat method, and citrate agar (CA) gel was used to stabilize O/W emulsions. The mechanisms of the CA structure and emulsion pH that affected emulsion stabilization were analyzed, and the application of CA gel emulsion (CAGE) was explored. Compared with native agar (NA), CA showed lower gel strength, higher transparency, and higher water contact angle. These changes indicate that a cross-linking reaction occurred, and it was demonstrated via FTIR and NMR. The emulsion properties were evaluated using particle size, ζ-potential, and the emulsification activity index. Results showed that CAGEs had a smaller particle size and lower ζ-potential than the native agar gel emulsion (NAGE). Meanwhile, confocal laser scanning microscopy confirmed that the CA gels stabilized the emulsions by forming a protective film around the oil droplets. Stability experiments revealed that CAGE (prepared with CA gel [DS = 0.145]) exhibited better stability than NAGE in the pH range of 3-11, and the rheological results further confirmed that the stability of the emulsions was influenced by the network structure and oil droplet interaction forces. Afterward, the application prospect of CAGE was evaluated by encapsulating vitamin D3 and curcumin.

Clinical Study of a Four-Step Program for the Treatment of Plantar Fasciitis with Bone Spurs.

OBJECTIVE: The most common causes of plantar and heel pain are plantar fasciitis and calcaneal spurs, and they often co-exist. Surgery is a recognized treatment for refractory plantar fasciitis. However, few studies have proposed treatment options for patients with metatarsophalangeal fasciitis with bone spurs. Accordingly, this study's purpose was to propose a four-step surgical regimen, and to improve the surgical outcome of plantar fasciitis with osteophytes and to establish a procedure for surgical treatment. METHODS: Retrospective analysis of 45 patients suffering from plantar fasciitis with bone spurs from 2020 to 2023. All patients underwent a four-step procedure, including plantar fascia release, calcaneal spur grinding, inflammatory tissue removal, and calcaneal burr decompression. The imaging parameters and functional scores were recorded before and after the operation. The objective evaluation included the measurement of calcaneal spur length on radiographs. Clinical evaluation included the American Orthopaedic Foot and Ankle Society (AOFAS), the Visual Analog Scale (VAS), and the Foot and Ankle Outcome Scale (FAOS). Measurement data that conformed to normal distribution were expressed as (x2 ± s), and pre-and postoperative AOFAS, FAOS, and VAS scores were compared using repeated-measures ANOVA, and preoperative and postoperative spur lengths were compared using paired t-tests. RESULTS: The 45 patients were followed up for 3 to 30 months, (17.72 ± 8.53) months, at final follow-up, the patient's AOFAS score improved from preoperative (74.93 ± 5.56) to (94.78 ± 3.98), FAOS score increased from preoperative (76.42 ± 3.37) to (96.16 ± 2.74), the VAS score decreased from (3.18 ± 0.54) to (1.07 ± 1.20) (p < 0.05), the length of spur decreased from (0.72 ± 1.81) cm to (0.23 ± 1.19) cm, and there were significant differences before and after operation (p < 0.05). CONCLUSION: The four-step surgical regimen is an appropriate and effective surgical procedure to treat plantar fasciitis with bone spurs.

Starch and sucrose metabolism plays an important role in the stem development in Medicago sativa.

The forage quality of alfalfa (Medicago sativa ) stems is greater than the leaves. Sucrose hydrolysis provides energy for stem development, with starch being enzymatically converted into sucrose to maintain energy homeostasis. To understand the physiological and molecular networks controlling stem development, morphological characteristics and transcriptome profiles in the stems of two alfalfa cultivars (Zhungeer and WL168) were investigated. Based on transcriptome data, we analysed starch and sugar contents, and enzyme activity related to starch-sugar interconversion. Zhungeer stems were shorter and sturdier than WL168, resulting in significantly higher mechanical strength. Transcriptome analysis showed that starch and sucrose metabolism were significant enriched in the differentially expressed genes of stems development in both cultivars. Genes encoding INV , bglX , HK , TPS and glgC downregulated with the development of stems, while the gene encoding was AMY upregulated. Weighted gene co-expression network analysis revealed that the gene encoding glgC was pivotal in determining the variations in starch and sucrose contents between the two cultivars. Soluble carbohydrate, sucrose, and starch content of WL168 were higher than Zhungeer. Enzyme activities related to sucrose synthesis and hydrolysis (INV, bglX, HK, TPS) showed a downward trend. The change trend of enzyme activity was consistent with gene expression. WL168 stems had higher carbohydrate content than Zhungeer, which accounted for more rapid growth and taller plants. WL168 formed hollow stems were formed during rapid growth, which may be related to the redistribution of carbohydrates in the pith tissue. These results indicated that starch and sucrose metabolism play important roles in the stem development in alfalfa.

Nomogram for predicting invasive lung adenocarcinoma in small solitary pulmonary nodules.

Background: This study aimed to construct a clinical prediction model and nomogram to differentiate invasive from non-invasive pulmonary adenocarcinoma in solitary pulmonary nodules (SPNs). Method: We analyzed computed tomography and clinical features as well as preoperative biomarkers in 1,106 patients with SPN who underwent pulmonary resection with definite pathology at Qilu Hospital of Shandong University between January 2020 and December 2021. Clinical parameters and imaging characteristics were analyzed using univariate and multivariate logistic regression analyses. Predictive models and nomograms were developed and their recognition abilities were evaluated using receiver operating characteristic (ROC) curves. The clinical utility of the nomogram was evaluated using decision curve analysis (DCA). Result: The final regression analysis selected age, carcinoembryonic antigen, bronchus sign, lobulation, pleural adhesion, maximum diameter, and the consolidation-to-tumor ratio as associated factors. The areas under the ROC curves were 0.844 (95% confidence interval [CI], 0.817-0.871) and 0.812 (95% CI, 0.766-0.857) for patients in the training and validation cohorts, respectively. The predictive model calibration curve revealed good calibration for both cohorts. The DCA results confirmed that the clinical prediction model was useful in clinical practice. Bias-corrected C-indices for the training and validation cohorts were 0.844 and 0.814, respectively. Conclusion: Our predictive model and nomogram might be useful for guiding clinical decisions regarding personalized surgical intervention and treatment options.

Nomogram combining clinical and radiological characteristics for predicting the malignant probability of solitary pulmonary nodules measuring ≤ 2 cm.

Background: At present, how to identify the benign or malignant nature of small (≤ 2 cm) solitary pulmonary nodules (SPN) are an urgent clinical challenge. This retrospective study aimed to develop a clinical prediction model combining clinical and radiological characteristics for assessing the probability of malignancy in SPNs measuring ≤ 2 cm. Method: In this study, we included patients with SPNs measuring ≤ 2 cm who underwent pulmonary resection with definite pathology at Qilu Hospital of Shandong University from January 2020 to December 2021. Clinical features, preoperative biomarker results, and computed tomography characteristics were collected. The enrolled patients were randomized at a ratio of 7:3 into a training cohort of 775 and a validation cohort of 331. The training cohort was used to construct the predictive model, while the validation cohort was used to test the model independently. Univariate and multivariate logistic regression analyses were performed to identify independent risk factors. The prediction model and nomogram were established based on the independent risk factors. The receiver operating characteristic (ROC) curve was used to evaluate the identification ability of the model. The calibration power was evaluated using the Hosmer-Lemeshow test and calibration curve. The clinical utility of the nomogram was also assessed by decision curve analysis (DCA). Result: A total of 1,106 patients were included in this study. Among them, the malignancy rate of SPNs was 85.08% (941/1,106). We finally identified the following six independent risk factors by logistic regression: age, carcinoembryonic antigen, nodule shape, calcification, maximum diameter, and consolidation-to-tumor ratio. The area under the ROC curve (AUC) for the training cohort was 0.764 (95% confidence interval [CI]: 0.714-0.814), and the AUC for the validation cohort was 0.729 (95% CI: 0.647-0.811), indicating that the prediction accuracy of nomogram was relatively good. The calibration curve of the predictive model also demonstrated a good calibration in both cohorts. DCA proved that the clinical prediction model was useful in clinical practice. Conclusion: We developed and validated a predictive model and nomogram for estimating the probability of malignancy in SPNs measuring ≤ 2 cm. With the application of predictive models, thoracic surgeons can make more rational clinical decisions while avoiding overtreatment and wasting medical resources.

Clinical characteristics and risk factors of lower extremity amputation in the diabetic inpatients with foot ulcers.

Objectives: To analyze clinical characteristics of the diabetic inpatients with foot ulcers and explore the risk factors of lower extremity amputation (LEA) in West China Hospital of Sichuan University. Methods: A retrospective analysis was performed based on the clinical data of the patients with diabetic foot ulcer (DFU) hospitalized in West China Hospital of Sichuan University from January 1, 2012 to December 31, 2020. The DFU patients were divided into three groups: non-amputation, minor amputation, and major amputation groups. The ordinal logistic regression analysis was used to identify the risk factors for LEA. Results: 992 diabetic patients (622 males and 370 females) with DFU were hospitalized in the Diabetic Foot Care Center of Sichuan University. Among them, 72 (7.3%) (55 minor amputations and 17 major amputations) cases experienced amputation, and 21(2.1%) refused amputation. Excluding the patients who refused amputation, the mean age and duration of diabetes of and HbA1c the 971 patients with DFU, were 65.1 ± 12.3 years old, 11.1 ± 7.6 years, and 8.6 ± 2.3% respectively. The patients in the major amputation group were older and had longer course of diabetes for a longer period of time than those in the non-amputation and minor amputation groups. Compared with the non-amputation patients (55.1%), more patients with amputation (minor amputation (63.5%) and major amputation (88.2%)) suffered from peripheral arterial disease (P=0.019). The amputated patients had statistically lower hemoglobin, serum albumin and ankle brachial index (ABI), but higher white blood cell, platelet counts, fibrinogen and C-reactive protein levels. The patients with amputation had a higher incidence of osteomyelitis (P = 0.006), foot gangrene (P < 0.001), and a history of prior amputations (P < 0.001) than those without amputation. Furthermore, a history of prior amputation (odds ratio 10.194; 95% CI, 2.646-39.279; P=0.001), foot gangrene (odds ratio 6.466; 95% CI, 1.576-26.539; P=0.010) and ABI (odds ratio 0.791; 95% CI, 0.639-0.980; P = 0.032) were significantly associated with LEAs. Conclusions: The DFU inpatients with amputation were older with long duration of diabetes, poorly glycemic control, malnutrition, PAD, severe foot ulcers with infection. A history of prior amputation, foot gangrene and a low ABI level were the independent predictors of LEA. Multidisciplinary intervention for DFU is essential to avoid amputation of the diabetic patients with foot ulcer.

mRNA therapeutics for disease therapy: principles, delivery, and clinical translation.

Messenger RNA (mRNA) has become a key focus in the development of therapeutic agents, showing significant potential in preventing and treating a wide range of diseases. The COVID-19 pandemic in 2020 has accelerated the development of mRNA nucleic therapeutics and attracted significant investment from global biopharmaceutical companies. These therapeutics deliver genetic information into cells without altering the host genome, making them a promising treatment option. However, their clinical applications have been limited by issues such as instability, inefficient in vivo delivery, and low translational efficiency. Recent advances in molecular design and nanotechnology have helped overcome these challenges, and several mRNA formulations have demonstrated promising results in both animal and human testing against infectious diseases and cancer. This review provides an overview of the latest research progress in structural optimization strategies and delivery systems, and discusses key considerations for their future clinical use.