Multifunctional hydrogels based on photothermal therapy: A prospective platform for the postoperative management of melanoma.

Preventing the recurrence of melanoma after surgery and accelerating wound healing are among the most challenging aspects of melanoma management. Photothermal therapy has been widely used to treat tumors and bacterial infections and promote wound healing. Owing to its efficacy and specificity, it may be used for postoperative management of tumors. However, its use is limited by the uncontrollable distribution of photosensitizers and the likelihood of damage to the surrounding normal tissue. Hydrogels provide a moist environment with strong biocompatibility and adhesion for wound healing owing to their highly hydrophilic three-dimensional network structure. In addition, these materials serve as excellent drug carriers for tumor treatment and wound healing. It is possible to combine the advantages of both of these agents through different loading modalities to provide a powerful platform for the prevention of tumor recurrence and wound healing. This review summarizes the design strategies, research progress and mechanism of action of hydrogels used in photothermal therapy and discusses their role in preventing tumor recurrence and accelerating wound healing. These findings provide valuable insights into the postoperative management of melanoma and may guide the development of promising multifunctional hydrogels for photothermal therapy.

Promoter hypomethylated PDZK1 acts as a tumorigenic gene in glioma by interacting with AKT1.

Glioma is the most frequently diagnosed primary brain tumor and typically has a poor prognosis because of malignant proliferation and invasion. It is urgent to elucidate the mechanisms driving glioma tumorigenesis and develop novel treatments to address this deadly disease. Here, we first revealed that PDZK1 is expressed at high levels in gliomas. Promoter hypomethylation may cause high expression of PDZK1 in glioma. Knockdown of PDZK1 inhibits glioma cell proliferation and invasion in vitro. Mechanistically, further investigations revealed that the loss of PDZK1 expression by siRNA inhibited the activation of the AKT/mTOR signaling pathway, leading to cell cycle arrest and apoptosis. Clinically, high expression of PDZK1 predicts a poorer prognosis for glioma patients than low expression of PDZK1. Overall, our study revealed that PDZK1 acts as a novel oncogene in glioma by binding to AKT1 and maintaining the activation of the AKT/mTOR signaling pathway. Thus, PDZK1 may be a potential therapeutic target for glioma.

Clinical and Dermoscopic Insights Into Scalp Nevi in Chinese Children.

BACKGROUND: The presence of scalp nevi in children frequently causes apprehension, leading physicians and parents to consider unnecessary biopsies or excisions of scalp nevi in children. There are limited data on the dermoscopic characteristics of scalp nevi in Chinese children. OBJECTIVE: The aim of this study was to comprehensively analyze the clinical and dermoscopic features of scalp nevi in this specific population of Chinese pediatric patients, with a focus on a single pediatric dermatologic surgery practice. METHODS: This retrospective cohort study investigated patients who underwent surgical excision of scalp nevi. All patients underwent dermoscopy with photographic documentation. RESULTS: Seventy-two scalp nevi in 56 Chinese children were included. Notably, no melanoma cases were detected. The parietal region (35, 48.6%) was the most frequently affected anatomical site. Clinical asymmetry was more prevalent in nevi with a diameter exceeding 6 mm (p < .05). The predominant dermoscopic pattern observed was the globular pattern (50, 69.4%) while an intriguing rarity of a reverse-eclipse pattern (1, 1.4%). CONCLUSION: This study revealed that scalp nevi in Chinese children usually did not exhibit concerning behavior. Increasing awareness of the clinical characteristics, dermoscopic features, and the natural progression of scalp nevi in children can potentially help reduce unnecessary surgical interventions.

[Relationship Between Tim-3 and Galectin-9 Expression Levels, Clinical Pathological Characteristics, and Prognosis in Patients After Radical Resection of Colorectal Cancer]. / ç»“ç›´è‚ ç™Œæ ¹æ²»æœ¯åŽæ‚£è€ Tim-3、galectin-9è¡¨è¾¾æ°´å¹³ä¸Žå ¶ä¸´åºŠç— ç†ç‰¹å¾åŠé¢„åŽçš„å ³ç³».

Objective: Some colorectal cancer patients still face high recurrence rates and poor prognoses even after they have undergone the surgical treatment of radical resection. Identifying potential biochemical markers and therapeutic targets for the prognostic evaluation of patients undergoing radical resection of colorectal cancer is crucial for improving their clinical outcomes. Recently, it has been reported that the T cell immunoglobulin and mucin domain protein 3 (Tim-3) and its ligand galactose lectin 9 (galectin-9) play crucial roles in immune dysfunction caused by various tumors, such as colorectal cancer. However, their expressions, biological functions, and prognostic value in colorectal cancer are still unclear. This study aims to investigate the relationship between Tim-3 and galectin-9 expression levels and the clinicopathological characteristics and prognosis of patients undergoing radical resection of colorectal cancer. Methods: A total of 171 patients who underwent radical resection of colorectal cancer at Chengdu Fifth People's Hospital between February 2018 and March 2019 were selected. Immunohistochemistry was performed to assess the expression levels of Tim-3 and galectin-9 in the cancer tissue samples and the paracancerous tissue samples of the patients. The relationship between Tim-3 and galectin-9 expression levels and the baseline clinical parameters of the patients was analyzed accordingly. Kaplan-Meier analysis was performed to assess the association between Tim-3 and galectin-9 expression levels and the relapse-free survival (RFS) and the overall survival (OS) of colorectal cancer patients. Cox regression analysis was conducted to identify factors associated with adverse prognosis in the patients. Results: The immunohistochemical results showed that the high expression levels of Tim-3 and galectin-9 were observed in 70.18% (120/171) and 32.16% (55/171), respectively, of the colorectal cancer tissues, whereas the low expression levels were 29.82% (51/171) and 67.84% (116/171), respectively. Furthermore, the expression score of Tim-3 was significantly higher in colorectal cancer tissues than that in the paracancerous tissues, while the expression score of galectin-9 was lower than that in the paracancerous tissues (P<0.05). Further analysis revealed that the expression of Tim-3 and galectin-9 was associated with the depth of tumor infiltration, vascular infiltration, and clinical staging (P<0.05). During the follow-up period of 14-63 months, 7 out of 171 patients were lost to follow-up. Among the remaining patients, 49 and 112 cases presented abnormally low expression of Tim-3 and galectin-9, respectively, whereas 115 and 52 cases presented high expression of Tim-3 and galectin-9, respectively. Kaplan-Meier survival analysis demonstrated that patients with high Tim-3 expression in colorectal cancer tissues had significantly lower RFS and OS than those with low expression did (RFS: log-rank=22.66, P<0.001; OS: log-rank=19.71, P<0.001). Conversely, patients with low galectin-9 expression had significantly lower RFS and OS than those with high expression did (RFS: log-rank=19.45, P<0.001; OS: log-rank=22.24, P<0.001). Cox multivariate analysis indicated that TNM stage Ⅲ (HR=2.26, 95% CI: 1.20-5.68), high expression of Tim-3 (HR=0.80, 95% CI: 0.33-0.91), and low expression of galectin-9 (HR=1.80, 95% CI: 1.33-4.70) were independent risk factors affecting RFS and OS in patients (P<0.05). Conclusion: Aberrant expression of Tim-3 and galectin-9 is observed in colorectal cancer tissues. High expression of Tim-3 and low expression of galectin-9 are closely associated with adverse clinico-pathological characteristics and prognosis. They are identified as independent influencing factors that may trigger adverse prognostic events in patients. These findings suggest that Tim-3 and galectin-9 have potential as new therapeutic targets and clinical indicators.

Reduced miR-99a-3p levels in systemic lupus erythematosus may promote B cell proliferation via NCAPG and the PI3K/AKT signaling pathway.

BACKGROUND: Systemic lupus erythematosus is an immunologically dysregulated disease characterized by the presence of multiple autoantibodies. In SLE, B lymphocytes contribute to the dysregulated production of autoantibodies and cytokines. Recently, we discovered that miR-99a-3p binds to both EIF4EBP1 and NCAPG mRNA and that lowering miR-99a-3p can promote B cell autophagy in SLE by increasing EIF4EBP1 expression. However, the functions of miR-99a-3p and NCAPG in SLE have not been extensively investigated. OBJECTIVE: This work aims to evaluate the levels of miR-99a-3p and NCAPG expression in SLE B cells and to determine whether the aberrant expression of miR-99a-3p and NCAPG contributes to the pathological mechanisms in SLE. METHODS: B lymphocytes were obtained through immunomagnetic negative selection. Using RT-qPCR, miR-99a-3p and NCAPG mRNA expressions in B lymphocytes and in the BALL-1 cell line were measured. To determine the relative abundance of NCAPG, PI3K, p-PI3K, AKT, and p-AKT, we normalize them to the level of ß-actin using Western blotting. Evaluation of miR-99a-3p and NCAPG's impact on cell proliferation was done utilizing CCK-8 assay. Using flow cytometry, the cell cycle and apoptosis were both measured. RESULTS: Comparing SLE B cells to healthy controls, miR-99a-3p expression was significantly downregulated. Additionally, it was observed that SLE B cells had significantly higher NCAPG mRNA expression. Blocking miR-99a-3p expression in BALL-1 cells with an antagomir elevated NCAPG expression, facilitated PI3K/AKT pathway activation, improved cell proliferation, raised the fraction of S-phase cells, and prevented cell apoptosis. The opposite effects of upregulated miR-99a-3p levels on BALL-1 cells were observed by using an agomir. Furthermore, the effect of decreased miR-99a-3p expression on cell proliferation was partially mediated by elevating NCAPG levels and activating the PI3K/AKT pathway. CONCLUSION: Our research indicates that lower miR-99a-3p expression in SLE B cells appears to boost B cell number via the NCAPG and PI3K/AKT pathways.

Functional characterization of group â ¡ interferon, IFNf in the acipenseriform fish, Chinese sturgeon (Acipenser sinensis).

Teleost fish possess a diversity of type Ⅰ interferons (IFNs) repertoire, which play a crucial role in antiviral and antimicrobial immune responses. In our previous study, IFNe1-3 and IFNb were identified and cloned from Chinese sturgeon (Acipenser sinensis), an acipenseriform fish. However, the absence of Chinese sturgeon genome data has left the question of whether there are other type Ⅰ IFN members in this species unresolved. In this study, we have identified and characterized a novel IFN, IFNf in Chinese sturgeon (AsIFNf). Bioinformatics analysis revealed that the AsIFNf contains a unique disulfide bond (2 cysteines) located in the second exon and fifth exon region, distinguishing it from other reported teleost type I IFNs. Meanwhile, qPCR results showed that AsIFNf mRNA was detectable in all examined tissues and up-regulated in the spleen or kidney in response to poly I: C, Citrobacter freundii, and Spring Viremia of Carp Virus (SVCV), but not by LPS. Furthermore, compared to recombinant AsIFNe2 protein (rAsIFNe2), rAsIFNf exhibited a stronger protective effect on Chinese sturgeon fin cells against SVCV and also induced higher expression of antiviral genes Mx and viperin. Importantly, AsIFNf displayed characteristics similar to antimicrobial peptides (AMPs) with a positive charge and demonstrated a broad spectrum of antimicrobial activity in vitro. These findings provide a theoretical foundation for understanding the primitive structure and function of interferon, as well as deepening our comprehension of the innate immune system and disease defense in the endangered Chinese sturgeon.

Extracellular vesicles from adipose stem cells ameliorate allergic rhinitis in mice by immunomodulatory.

Background: Human adipose tissue-derived stem cells (hADSCs) exert potent immunosuppressive effects in the allogeneic transplantation treatment. In mouse model of allergic rhinitis (AR), ADSCs partially ameliorated AR. However, no study has evaluated the potential therapeutic effects of hADSC-derived extracellular vesicles (hADSC-EVs) on AR. Methods: Female BALB/c mice were sensitized and challenged with ovalbumin (OVA) to induce AR. One day after the last nasal drop, each group received phosphate buffered saline (PBS) or hADSC-EVs treatment. Associated symptoms and biological changes were then assessed. Results: hADSC-EV treatment significantly alleviated nasal symptoms, and reduced inflammatory infiltration. Serum levels of OVA-specific IgE, interleukin (IL)-4 and interferon (IFN)-γ were all significantly reduced. The mRNA levels of IL-4 and IFN-γ in the spleen also changed accordingly. The T helper (Th)1/Th2 cell ratio increased. The treatment efficacy index of hADSC-EV was higher than that of all human-derived MSCs in published reports on MSC treatment of AR. ADSC-EVs exhibited a greater therapeutic index in most measures when compared to our previous treatment involving ADSCs. Conclusion: These results demonstrated that hADSC-EVs could ameliorate the symptoms of AR by modulating cytokine secretion and Th1/Th2 cell balance. hADSC-EVs could potentially be a viable therapeutic strategy for AR. Further animal studies are needed to elucidate the underlying mechanisms and to optimize potential clinical protocols.

Diagnosis and surgical outcomes of coarctation of the aorta in pediatric patients: a retrospective study.

Background: Coarctation of the aorta (CoA) is a common congenital cardiovascular malformation, and improvements in the diagnostic process for surgical decision-making are important. We sought to compare the diagnostic accuracy of transthoracic echocardiography (TTE) with computed tomographic angiography (CTA) to diagnose CoA. Methods: We retrospectively reviewed 197 cases of CoA diagnosed by TTE and CTA and confirmed at surgery from July 2009 to August 2019. Results: The surgical findings confirmed that 19 patients (9.6%) had isolated CoA and 178 (90.4%) had CoA combined with other congenital cardiovascular malformations. The diagnostic accuracy of CoA by CTA was significantly higher than that of TTE (χ2 = 6.52, p = 0.01). In contrast, the diagnostic accuracy of TTE for associated cardiovascular malformations of CoA was significantly higher than that of CTA (χ2 = 15.36, p < 0.0001). Infants and young children had more preductal type of CoA, and PDA was the most frequent cardiovascular lesion associated with CoA. The pressure gradient was significantly decreased after the first operation, similar at 6 months, 1 year, and 3 years follow-ups by TTE. Conclusions: CTA is more accurate as a clinical tool for diagnosing CoA; however, TTE with color Doppler can better identify associated congenital cardiovascular malformations. Therefore, combining TTE and CTA would benefit clinical evaluation and management in patients suspected of CoA. TTE was valuable for post-operation follow-up and clinical management.

Genetic and phenotypic diversities of nevus spilus phenotypes: Case series and a proposed diagnostic algorithm.

Nevus spilus (NS) is composed of multiple types that characterized by a congenital hyperpigmented patch within variable even superimposed lesions originating from melanocytic lineage cells. The molecular mechanism and classification of diverse NS phenotypes remain unclear. Five children with a phenotype of NS were genotyped by the panel based on next-generation sequencing in this study. DNA from biopsies, blood samples and hair follicle were sequenced to confirm the presence of a somatic mutation. Sequencing results indicated somatic mutation in the gene of NRAS or HRAS in all biopsies from the nevi, and the pathogenic variants were not detected in the samples of blood and hair follicle. This study successfully identified the somatic mutation in five unrelated children with diverse NS phenotypes. Moreover, it provided typical images and differential diagnoses between variable NS phenotypes in clinical, pathological, and genetic features, and first proposed a clinical diagnostic algorithm that contributed to simplifying and optimizing the diagnoses and management of these overlapped diseases.

[Corrigendum] Roles of the H19/microRNA­675 axis in the proliferation and epithelial­mesenchymal transition of human cutaneous squamous cell carcinoma cells.

Following the publication of the above paper, a concerned reader drew to the authors' attention that, in Fig. 4C on p. 8, the 'Invasion, miR­675­inhibitor' data panel appeared to contain an overlapping section with the 'Invasion, mi­R675­inhibitor + pcDNA3.1­H19' data panel for the SCL1 cell line, such that the data were likely to have been derived from the same original source, even though they were intended to show the results from differently performed experiments. After having examined the original data, the authors also realized that the 'Inhibitor­NC' and 'miR­675­inhibitor' data panels showing the migration assay experiments for the A431 cell line in the same figure part had also inadvertently been derived from the same original source. After having been granted permission from the Editor of Oncology Reports to repeat the experiments shown in Fig. 4C, the revised version of Fig. 4, incorporating the new data for Fig. 4C, is shown on the next page. Note that these errors did not affect the overall conclusions reported in the study, and the repeated experiment yielded similar results to those obtained originally. The authors are grateful to the Editor for allowing them the opportunity to publish this corrigendum, and all the authors agree with the publication of this; furthermore, they apologize for any inconvenience caused to the readership of the Journal. [Oncology Reports 45: 39, 2021; DOI: 10.3892/or.2021.7990].

Prenylated PALM2 Promotes the Migration of Esophageal Squamous Cell Carcinoma Cells Through Activating Ezrin.

Proteins containing a CAAX motif at the C-terminus undergo prenylation for localization and activity and include a series of key regulatory proteins, such as RAS superfamily members, heterotrimeric G proteins, nuclear lamina protein, and several protein kinases and phosphatases. However, studies of prenylated proteins in esophageal cancer are limited. Here, through research on large-scale proteomic data of esophageal cancer in our laboratory, we found that paralemmin-2 (PALM2), a potential prenylated protein, was upregulated and associated with poor prognosis in patients. Low-throughput verification showed that the expression of PALM2 in esophageal cancer tissues was higher than that in their paired normal esophageal epithelial tissues, and it was generally expressed in the membrane and cytoplasm of esophageal cancer cells. PALM2 interacted with the two subunits of farnesyl transferase (FTase), FNTA and FNTB. Either the addition of an FTase inhibitor or mutation in the CAAX motif of PALM2 (PALM2C408S) impaired its membranous localization and reduced the membrane location of PALM2, indicating PALM2 was prenylated by FTase. Overexpression of PALM2 enhanced the migration of esophageal squamous cell carcinoma cells, whereas PALM2C408S lost this ability. Mechanistically, PALM2 interacted with the N-terminal FERM domain of ezrin of the ezrin/radixin/moesin (ERM) family. Mutagenesis indicated that lysine residues K253/K254/K262/K263 in ezrin's FERM domain and C408 in PALM2's CAAX motif were important for PALM2/ezrin interaction and ezrin activation. Knockout of ezrin prevented enhanced cancer cell migration by PALM2 overexpression. PALM2, depending on its prenylation, increased both ezrin membrane localization and phosphorylation of ezrin at Y146. In summary, prenylated PALM2 enhances the migration of cancer cells by activating ezrin.

Co-Expression of Chromatin Assembly Factor 1 Subunit A and Proliferating Cell Nuclear Antigen Is a Prognostic Biomarker of Esophageal Cancer.

(1) Background: Esophageal cancer (EC) is an important global health challenge. Due to the lack of necessary biomarkers and therapeutic targets, the survival of EC patients is poor. The EC proteomic data of 124 patients recently published by our group provides a database for research in this field. (2) Methods: Bioinformatics analysis was used to identify DNA replication and repair-related proteins in EC. Proximity ligation assay, colony formation assay, DNA fiber assay, and flow cytometry were used to study the effects of related proteins on EC cells. Kaplan-Meier survival analysis was used to evaluate the relationship between gene expression and the survival time of EC patients. (3) Results: Chromatin assembly factor 1 subunit A (CHAF1A) was highly correlated with proliferating cell nuclear antigen (PCNA) expression in EC. CHAF1A and PCNA colocalized in the nucleus of EC cells. Compared with the knockdown of CHAF1A or PCNA alone, the double knockdown of CHAF1A and PCNA could significantly inhibit EC cell proliferation. Mechanistically, CHAF1A and PCNA synergistically accelerated DNA replication and promoted S-phase progression. EC patients with high expression of both CHAF1A and PCNA had a worse survival rate. (4) Conclusion: we identify CHAF1A and PCNA as key cell cycle-related proteins leading to the malignant progression of EC, and these proteins could serve as important prognostic biomarkers and targets for EC.

Sulconazole Induces PANoptosis by Triggering Oxidative Stress and Inhibiting Glycolysis to Increase Radiosensitivity in Esophageal Cancer.

Esophageal cancer is the seventh most common cancer in the world. Although traditional treatment methods such as radiotherapy and chemotherapy have good effects, their side effects and drug resistance remain problematic. The repositioning of drug function provides new ideas for the research and development of anticancer drugs. We previously showed that the Food and Drug Administration-approved drug sulconazole can effectively inhibit the growth of esophageal cancer cells, but its molecular mechanism is not clear. Here, our study demonstrated that sulconazole had a broad spectrum of anticancer effects. It can not only inhibit the proliferation but also inhibit the migration of esophageal cancer cells. Both transcriptomic sequencing and proteomic sequencing showed that sulconazole could promote various types of programmed cell death and inhibit glycolysis and its related pathways. Experimentally, we found that sulconazole induced apoptosis, pyroptosis, necroptosis, and ferroptosis. Mechanistically, sulconazole triggered mitochondrial oxidative stress and inhibited glycolysis. Finally, we showed that low-dose sulconazole can increase radiosensitivity of esophageal cancer cells. Taken together, these new findings provide strong laboratory evidence for the clinical application of sulconazole in esophageal cancer.

The function, mechanisms, and clinical applications of metformin: potential drug, unlimited potentials.

Metformin has been used clinically for more than 60 years. As time goes by, more and more miraculous effects of metformin beyond the clinic have been discovered and discussed. In addition to the clinically approved hypoglycemic effect, it also has a positive metabolic regulation effect on the human body that cannot be ignored. Such as anti-cancer, anti-aging, brain repair, cardiovascular protection, gastrointestinal regulation, hair growth and inhibition of thyroid nodules, and other nonclinical effects. Metformin affects almost the entire body in the situation taking it over a long period, and the preventive effects of metformin in addition to treating diabetes are also beginning to be recommended in some guidelines. This review is mainly composed of four parts: the development history of metformin, the progress of clinical efficacy, the nonclinical efficacy of metformin, and the consideration and prospect of its application.

Circ_0078607 increases platinum drug sensitivity via miR-196b-5p/GAS7 axis in ovarian cancer.

Platinum-based chemotherapy is one of the predominant strategies for treating ovarian cancer (OC), however, platinum resistance greatly influences the therapeutic effect. Circular RNAs (circRNAs) have been found to participate in the pathogenesis of platinum resistance. Our aim was to explore the involvement of circ_0078607 in OC cell cisplatin (DDP) resistance and its potential mechanisms. Circ_0078607, miR-196b-5p, and growth arrest-specific 7 (GAS7) levels were assessed by qPCR. Circ_0078607 stability was assessed by ribonuclease R digestion and actinomycin D treatment. Cell viability of various conic of DDP treatment was measured by CCK-8. The cell proliferation was determined by CCK-8 and colony formation assay. Western blotting was performed for determining GAS7, ABCB1, CyclinD1 and Bcl-2 protein levels. The direct binding between miR-196b-5p and circ_0078607 or GAS7 was validated by dual-luciferase reporter and RIP assay. DDP resistance in vivo was evaluated in nude mice. Immunohistochemistry staining for detecting Ki67 expression in xenograft tumours. Circ_0078607 and GAS7 was down-regulated, but miR-196b-5p was up-regulated in OC samples and DDP-resistant cells. Overexpression of circ_0078607 inhibited DDP resistance, cell growth and induced apoptosis in DDP-resistant OC cells. Mechanistically, circ_0078607 sequestered miR-196b-5p to up-regulate GAS7. MiR-196b-5p mimics reversed circ_0078607 or GAS7 overexpression-mediated enhanced sensitivity. Finally, circ_0078607 improved the sensitivity of DDP in vivo. Circ_0078607 attenuates DDP resistance via miR-196b-5p/GAS7 axis, which highlights the therapeutic potential of circ_0078607 to counter DDP resistance in OC.

Research Progress of Traditional Chinese Medicine Monomers in Reversing Multidrug Resistance of Breast Cancer.

Breast cancer is a malignant disease with an increasing incidence. Chemotherapy is still an important means for breast cancer treatment, but multidrug resistance (MDR) greatly limits its clinical application. Therefore, the high-efficiency MDR reversal agents are urgently needed. Traditional Chinese medicine (TCM) monomers have unique advantages in reversing chemotherapeutic MDR because of its low toxicity, high efficiency, and ability to impact multiple targets. This review firstly summarizes the major mechanisms of MDR in breast cancer, including the reduced accumulation of intracellular chemotherapeutic drugs, the promoted inactivation of intracellular chemotherapeutic drugs, and the enhanced damage repair ability of DNA, etc., and secondly highlights the research progress of 15 kinds of TCM monomers, including curcumin, resveratrol, emodin, apigenin, tetrandrine, gambogic acid, matrine, paeonol, schisandrin B, [Formula: see text]-elemene, astragaloside IV, berberine, puerarin, tanshinone IIA, and quercetin, in reversing MDR of breast cancer. This review also provides the suggestion for the future research of MDR reversal agents in breast cancer.

Effect of Cooking Method and Doneness Degree on Volatile Compounds and Taste Substance of Pingliang Red Beef.

This study used gas chromatography-ion mobility spectrometry (GC-IMS) and high-performance liquid chromatography (HPLC) methods to examine the impact of cooking methods and doneness on volatile aroma compounds and non-volatile substances (fatty acids, nucleotides, and amino acids) in Pingliang red beef. The flavor substances' topographic fingerprints were established, and 45 compounds were traced to 71 distinct signal peaks. Pingliang red beef's fruity flavor was enhanced thanks to the increased concentration of hexanal, styrene, and 2-butanone that resulted from instant boiling. The levels of 3-methylbutanal, which contributes to the characteristic caramel-chocolate-cheese aroma, peaked at 90 min of boiling and 40 min of roasting. The FFA content was reduced by 28.34% and 27.42%, respectively, after the beef was roasted for 40 min and instantly boiled for 10 s (p > 0.05). The most distinctive feature after 30 min of boiling was the umami, as the highest levels of glutamate (Glu) (p < 0.05) and the highest equivalent umami concentration (EUC) values were obtained through this cooking method. Additionally, adenosine-5'-monophosphate (AMP) and inosine-5'-monophosphate (IMP) decreased with increasing doneness compared to higher doneness, indicating that lower doneness was favorable in enhancing the umami of the beef. In summary, different cooking methods and doneness levels can affect the flavor and taste of Pingliang red beef, but it is not suitable for high-doneness cooking.