A qualitative transcriptional signature for the histological reclassification of lung squamous cell carcinomas and adenocarcinomas.

BACKGROUND: Targeted therapy for non-small cell lung cancer is histology dependent. However, histological classification by routine pathological assessment with hematoxylin-eosin staining and immunostaining for poorly differentiated tumors, particularly those from small biopsies, is still challenging. Additionally, the effectiveness of immunomarkers is limited by technical inconsistencies of immunostaining and lack of standardization for staining interpretation. RESULTS: Using gene expression profiles of pathologically-determined lung adenocarcinomas and squamous cell carcinomas, denoted as pADC and pSCC respectively, we developed a qualitative transcriptional signature, based on the within-sample relative gene expression orderings (REOs) of gene pairs, to distinguish ADC from SCC. The signature consists of two genes, KRT5 and AGR2, which has the stable REO pattern of KRT5 > AGR2 in pSCC and KRT5 < AGR2 in pADC. In the two test datasets with relative unambiguous NSCLC types, the apparent accuracy of the signature were 94.44 and 98.41%, respectively. In the other integrated dataset for frozen tissues, the signature reclassified 4.22% of the 805 pADC patients as SCC and 12% of the 125 pSCC patients as ADC. Similar results were observed in the clinical challenging cases, including FFPE specimens, mixed tumors, small biopsy specimens and poorly differentiated specimens. The survival analyses showed that the pADC patients reclassified as SCC had significantly shorter overall survival than the signature-confirmed pADC patients (log-rank p = 0.0123, HR = 1.89), consisting with the knowledge that SCC patients suffer poor prognoses than ADC patients. The proliferative activity, subtype-specific marker genes and consensus clustering analyses also supported the correctness of our signature. CONCLUSIONS: The non-subjective qualitative REOs signature could effectively distinguish ADC from SCC, which would be an auxiliary test for the pathological assessment of the ambiguous cases.

An individualised signature for predicting response with concordant survival benefit for lung adenocarcinoma patients receiving platinum-based chemotherapy.

BACKGROUND: For lung adenocarcinoma (LUAD) patients receiving platinum-based adjuvant chemotherapy (ACT), predictive signatures extracted from survival data solely are not directly associated with platinum response. Another limitation of reported signatures, commonly based on risk scores summarised from gene expressions, is that they could not be applied directly to samples measured by different laboratories due to experimental batch effects. METHODS: Using 60 samples of LUAD patients receiving platinum-based ACT in TCGA, we pre-selected gene pairs whose within-samples relative expression orderings (REOs) were significantly associated with both pathological response and 5-year survival, from which we selected an optimal signature whose within-samples REOs could identify responders with improved 5-year survival rate. RESULTS: A predictive signature consisting of three gene pairs was developed. In an independent data set integrated from five small data sets, the predicted responders had a significantly higher 5-year survival rate than the predicted non-responders if and only if they received platinum-based ACT (log-rank P=0.0006). The predicted responders showed a 22% absolute benefit of platinum-based ACT in 5-year survival rate compared with untreated patients (log-rank P=0.0019). CONCLUSIONS: The REO-based signature can individually predict response to platinum-based ACT with concordant survival benefit directly for LUAD samples measured by different laboratories.

Hepatoprotective effect and structural analysis of Hedysarum polysaccharides in vivo and in vitro.

The crude Hedysarum polysaccharides (HPS: HPS-50 and HPS-80) obtained from Radix Hedysari exhibited great pharmacological activities in our previous research. This study investigated the effects of HPS on lipopolysaccharide (LPS)/D-galactosamine (D-GalN)-induced acute liver injury (ALI) in mice and LPS-induced injury in LO2 cells, as well as the relationship between structural characteristics and hepatoprotective activities. The in vivo results showed that compared with HPS-80, HPS-50 showed stronger hepatoprotection, which improved histopathological changes to normal levels. HPS-50 significantly decreased the levels of ALT, AST, MPO, and MDA, increased the activities of SOD, CAT, and GSH, and suppressed the LPS/D-GalN-triggered production of TNF-α, IL-1ß, and IL-6 (p < .05). The results in vitro showed that HPS-50-P (HPS-50-1, HPS-50-2, and HPS-50-3) purified from HPS-50 played significant protective roles against LPS-induced injury in LO2 cells by reducing cell apoptosis and relieving cell cycle arrest. HPS-50-2 restored the percentage of normal cells from 54.8% to 94.7%, and reduced the S phase cells from 59.40% to 47.05% (p < .01). By analyzing the structure of HPS-50-P, including monosaccharide composition, molecular weight, chain conformation, and surface morphology, we speculated that the best protective effect of HPS-50-2 might be attributed to its beta configuration, highest molecular weight, and high glucose and galactose contents. These findings indicate that HPS-50 might be a promising source of functional foods for the protection and prevention of ALI. PRACTICAL APPLICATIONS: In this study, the protective effect of HPS on ALI was evaluated from multiple perspectives, and HPS-50-2 was screened as a potential active ingredient. This study has two practical applications. First, it provides a new way to improve ALI, and a new option for patients to prevent and treat ALI. Second, this work also complements the pharmacological activity of Radix Hedysari and provides a basis for the development of Radix Hedysari as a functional food.

Correction: Formononetin in Radix Hedysari extract-mediated green synthesis of gold nanoparticles for colorimetric detection of ferrous ions in tap water.

[This corrects the article DOI: 10.1039/D0RA05660J.].

The amelioration of ulcerative colitis induced by Dinitrobenzenesulfonic acid with Radix Hedysari.

Ulcerative colitis (UC) is a chronic inflammatory disease with an unknown precise etiology. This study proves that Radix Hedysari (RH) ameliorates UC. Four RH extracts were used to ameliorate UC induced by 2,4-Dinitrobenzenesulfonic acid by 7 days intervention in agreement to preliminary studies. Compared to treatment with RH extracts, the RH ethanol extract (EE) was found to be more effective in ameliorating UC. With EE, the DAI were significantly decreased. Macroscopic and histopathological assessments suggest that the colon mucosa was repaired, the organizational structure of the colon had been rebuilt. The levels of MPO, TNF-α, IL-1ß, and MDA were significantly decreased (p < .01), the levels of T-SOD and CAT were significantly increased (p < .01). Moreover, the compounds in EE were analyzed by HPLC. The results show that EE can ameliorate UC, and its anti-inflammatory capability probably plays an important role. RH can act as a functional food and ameliorate UC. PRACTICAL APPLICATIONS: In this work, the ameliorative effect of RH on UC was evaluated from multiple angles. There are two practical applications of this work. On the one hand, a new approach to ameliorating UC is provided by this work. In addition, UC patients have a new option for improving their symptoms. On the other hand, this work also provides information on how best to process RH for therapeutic use. In addition, we can utilize some compounds of RH that were once considered useless and reduce the waste of natural resources.

Formononetin in Radix Hedysari extract-mediated green synthesis of gold nanoparticles for colorimetric detection of ferrous ions in tap water.

This study linked natural plant materials and nanomaterials; reporting an environmentally friendly, non-toxic and efficient method for the green synthesis of gold nanoparticles (AuNPs) using an ethyl acetate extract of Radix Hedysari (EAR). The components of the extract were identified using HPLC and it was found that formononetin accounted for more than 90% of the total contents. We predicted that formononetin in EAR plays a crucial role in green synthesis. Thus, formononetin was used as a standard reductant to synthesize AuNPs, and the result confirmed our prediction. The synthetic mechanism was also discussed in detail in the article. Moreover, EAR-AuNPs realized the sensitive and selective colorimetric detection of ferrous ions (Fe2+) among other metal ions, and were applied to spiked tap water with a low detection limit of 1.5 µM in a wide range from 10 µM to 500 µM. EAR-AuNPs were green synthesized using Radix Hedysari extract for the first time and were successfully applied in real sample detection.

Protective effect of polysaccharides from Radix Hedysari on gastric ulcers induced by acetic acid in rats.

The dry root of Hedysarum polybotrys Hand.-Mazz., commonly known as "Hong Qi", has a variety of health benefits. The present study was undertaken to explore the anti-gastric ulcer potential effect of Hedysarum polysaccharides (HPS; HPS-50, HPS-80), the principal active fraction of Radix Hedysari (RH). The anti-gastric ulcer effects of HPS were evaluated using an animal model of ulcerative lesions induced by acetic acid. The effects of antioxidant factors, anti-inflammatory cytokines, and mucosal blood flow regulatory factor levels in the gastric tissue homogenate of rats were analyzed for the bioactivities of HPS. The results showed that, compared with the acetic acid-induced ulcerated group, the ulcer inhibition rate of HPS-treated rats was significantly increased. The pathological findings suggested that mucosal regeneration, cell migration, and inflammatory cell infiltration were decreased, and collagen fibers were significantly reduced. Extensive granulation tissue proliferation indicated the healing stage was initiated, suggesting a good prognosis. The oxidative stress status of the gastric ulcer rats was improved, the levels of TNF-α and IL-6 were significantly decreased, and the levels of PGE-2 and NO were increased (P < 0.05). HPS-80-H may be a promising ingredient for incorporation into functional foods or nutritional supplements for the prevention of gastric ulcers.

Structure-immunomodulatory activity relationships of Hedysarum polysaccharides extracted by a method involving a complex enzyme combined with ultrasonication.

A new, more effective and environmentally friendly method involving a complex enzyme combined with ultrasonication was employed to extract and isolate three novel polysaccharides (HPS-MCs: HPS-MC, HPS-MC (50%) and HPS-MC (80%)) of Radix Hedysari. Compared with polysaccharides obtained using a traditional extraction method (hot water extraction, HPS-R), the yields and total carbohydrate contents of HPS-MCs were significantly higher. HPS-MC (80%) exhibited relatively strong immunomodulatory activity and a concentration-dependent dose-response relationship under cyclophosphamide (CP)-induced immunosuppressive conditions in mice models. To more comprehensively investigate the relationships between structural characteristics and immunomodulatory activity, HPS-MC (80%) was fractionated into three major homogeneous polysaccharide fractions (HPS-MC (80%)s: HPS-MC (80%)-1, HPS-MC (80%)-2, and HPS-MC (80%)-3). These three homogeneous polysaccharides had different mass percentages of monosaccharides species (rhamnose, arabinose, mannose, glucose, and galactose) by gas chromatography (GC) and different molecular weights and chain conformations by high-performance gel permeation chromatography coupled with multi-angle laser light scattering (HPGPC-MALLS), and promoted macrophage and splenocyte proliferation to different degrees. These findings indicated that HPS-MC (80%) had a prominent potential immune response, especially HPS-MC (80%)-2 and HPS-MC (80%)-3, and might be suitable candidates for functional foods or potential novel immunomodulators.

An individualized gene expression signature for prediction of lung adenocarcinoma metastases.

Our laboratory previously reported an individual-level signature consisting of nine gene pairs, named 9-GPS. This signature was developed by training on microarray expression data and validated using three independent integrated microarray data sets, with samples of stage I non-small-cell lung cancer after complete surgical resection. In this study, we first validated the cross-platform robustness of 9-GPS by demonstrating that 9-GPS could significantly stratify the overall survival of 213 stage I lung adenocarcinoma (LUAD) patients detected with RNA-sequencing platform in The Cancer Genome Atlas (TCGA; log-rank P = 0.0318, C-index = 0.55). Applying 9-GPS to all the 423 stage I-IV LUAD samples in TCGA, the predicted high-risk samples were significantly enriched with clinically diagnosed metastatic samples (Fisher's exact test, P = 0.0015). We further modified the voting rule of 9-GPS and found that the modified 9-GPS had a better performance in predicting metastasis states (Fisher's exact test, P < 0.0001). With the aid of the modified 9-GPS for reclassifying the metastasis states of patients with LUAD, the reclassified metastatic samples presented clearer transcriptional and genomic characteristics compared to the reclassified nonmetastatic samples. Finally, regulator network analysis identified TP53 and IRF1 with frequent genomic aberrations in the reclassified metastatic samples, indicating their key roles in driving tumor metastasis. In conclusion, 9-GPS is a robust signature for identifying early-stage LUAD patients with potential occult metastasis. This occult metastasis prediction was associated with clear transcriptional and genomic characteristics as well as the clinical diagnoses.