Uncharacterized yeast gene YBR238C, an effector of TORC1 signaling in a mitochondrial feedback loop, accelerates cellular aging via HAP4- and RMD9-dependent mechanisms.

Uncovering the regulators of cellular aging will unravel the complexity of aging biology and identify potential therapeutic interventions to delay the onset and progress of chronic, aging-related diseases. In this work, we systematically compared genesets involved in regulating the lifespan of Saccharomyces cerevisiae (a powerful model organism to study the cellular aging of humans) and those with expression changes under rapamycin treatment. Among the functionally uncharacterized genes in the overlap set, YBR238C stood out as the only one downregulated by rapamycin and with an increased chronological and replicative lifespan upon deletion. We show that YBR238C and its paralog RMD9 oppositely affect mitochondria and aging. YBR238C deletion increases the cellular lifespan by enhancing mitochondrial function. Its overexpression accelerates cellular aging via mitochondrial dysfunction. We find that the phenotypic effect of YBR238C is largely explained by HAP4- and RMD9-dependent mechanisms. Furthermore, we find that genetic- or chemical-based induction of mitochondrial dysfunction increases TORC1 (Target of Rapamycin Complex 1) activity that, subsequently, accelerates cellular aging. Notably, TORC1 inhibition by rapamycin (or deletion of YBR238C) improves the shortened lifespan under these mitochondrial dysfunction conditions in yeast and human cells. The growth of mutant cells (a proxy of TORC1 activity) with enhanced mitochondrial function is sensitive to rapamycin whereas the growth of defective mitochondrial mutants is largely resistant to rapamycin compared to wild type. Our findings demonstrate a feedback loop between TORC1 and mitochondria (the TORC1-MItochondria-TORC1 (TOMITO) signaling process) that regulates cellular aging processes. Hereby, YBR238C is an effector of TORC1 modulating mitochondrial function.

Tanshinone IIA induces ER stress and JNK activation to inhibit tumor growth and enhance anti-PD-1 immunotherapy in non-small cell lung cancer.

BACKGROUND: Non-small cell lung cancer (NSCLC) remains at the forefront of new cancer cases, and there is an urgent need to find new treatments or improve the efficacy of existing therapies. In addition to the application in the field of cerebrovascular diseases, recent studies have revealed that tanshinone IIA (Tan IIA) has anticancer activity in a variety of cancers. PURPOSE: To investigate the potential anticancer mechanism of Tan IIA and its impact on immunotherapy in NSCLC. METHODS: Cytotoxicity and colony formation assays were used to detect the Tan IIA inhibitory effect on NSCLC cells. This research clarified the mechanisms of Tan IIA in anti-tumor and programmed death-ligand 1 (PD-L1) regulation by using flow cytometry, transient transfection, western blotting and immunohistochemistry (IHC) methods. Besides, IHC was also used to analyze the nuclear factor of activated T cells 1 (NFAT2) expression in NSCLC clinical samples. Two animal models including xenograft mouse model and Lewis lung cancer model were used for evaluating tumor suppressive efficacy of Tan IIA. We also tested the efficacy of Tan IIA combined with programmed cell death protein 1 (PD-1) inhibitors in Lewis lung cancer model. RESULTS: Tan IIA exhibited good NSCLC inhibitory effect which was accompanied by endoplasmic reticulum (ER) stress response and increasing Ca2+ levels. Moreover, Tan IIA could suppress the NFAT2/ Myc proto oncogene protein (c-Myc) signaling, and it also was able to control the Jun Proto-Oncogene(c-Jun)/PD-L1 axis in NSCLC cells through the c-Jun N-terminal kinase (JNK) pathway. High NFAT2 levels were potential factors for poor prognosis in NSCLC patients. Finally, animal experiments data showed a stronger immune activation phenotype, when we performed treatment of Tan IIA combined with PD-1 monoclonal antibody. CONCLUSION: The findings of our research suggested a novel mechanism for Tan IIA to inhibit NSCLC, which could exert anti-cancer effects through the JNK/NFAT2/c-Myc pathway. Furthermore, Tan IIA could regulate tumor PD-L1 levels and has the potential to improve the efficacy of PD-1 inhibitors.

Clinical Evaluation of the Effect of Encorafenib on Bupropion, Rosuvastatin, and Coproporphyrin I and Considerations for Statin Coadministration.

BACKGROUND AND OBJECTIVES: Encorafenib is a kinase inhibitor indicated for the treatment of patients with unresectable or metastatic melanoma or metastatic colorectal cancer, respectively, with selected BRAF V600 mutations. A clinical drug-drug interaction (DDI) study was designed to evaluate the effect of encorafenib on rosuvastatin, a sensitive substrate of OATP1B1/3 and breast cancer resistance protein (BCRP), and bupropion, a sensitive CYP2B6 substrate. Coproporphyrin I (CP-I), an endogenous substrate for OATP1B1, was measured in a separate study to deconvolute the mechanism of transporter DDI. METHODS: DDI study participants received a single oral dose of rosuvastatin (10 mg) and bupropion (75 mg) on days - 7, 1, and 14 and continuous doses of encorafenib (450 mg QD) and binimetinib (45 mg BID) starting on day 1. The CP-I data were collected from participants in a phase 3 study who received encorafenib (300 mg QD) and cetuximab (400 mg/m2 initial dose, then 250 mg/m2 QW). Pharmacokinetic and pharmacodynamic analysis was performed using noncompartmental and compartmental methods. RESULTS: Bupropion exposure was not increased, whereas rosuvastatin Cmax and area under the receiver operating characteristic curve (AUC) increased approximately 2.7 and 1.6-fold, respectively, following repeated doses of encorafenib and binimetinib. Increase in CP-I was minimal, suggesting that the primary effect of encorafenib on rosuvastatin is through BCRP. Categorization of statins on the basis of their metabolic and transporter profile suggests pravastatin would have the least potential for interaction when coadministered with encorafenib. CONCLUSION: The results from these clinical studies suggest that encorafenib does not cause clinically relevant CYP2B6 induction or inhibition but is an inhibitor of BCRP and may also inhibit OATP1B1/3 to a lesser extent. Based on these results, it may be necessary to consider switching statins or reducing statin dosage accordingly for coadministration with encorafenib. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov NCT03864042, registered 6 March 2019.

PICLS with human cells is the first high throughput screening method for identifying novel compounds that extend lifespan.

Gerontology research on anti-aging interventions with drugs could be an answer to age-related diseases, aiming at closing the gap between lifespan and healthspan. Here, we present two methods for assaying chronological lifespan in human cells: (1) a version of the classical outgrowth assay with quantitative assessment of surviving cells and (2) a version of the PICLS method (propidium iodide fluorescent-based measurement of cell death). Both methods are fast, simple to conduct, cost-effective, produce quantitative data for further analysis and can be used with diverse human cell lines. Whereas the first method is ideal for validation and testing the post-intervention reproductive potential of surviving cells, the second method has true high-throughput screening potential. The new technologies were validated with known anti-aging compounds (2,5-anhydro-D-mannitol and rapamycin). Using the high-throughput screening method, we screened a library of 162 chemical entities and identified three compounds that extend the longevity of human cells.

Identification of heterosis and combining ability in the hybrids of male sterile and restorer sorghum [Sorghum bicolor (L.) Moench] lines.

In sorghum [Sorghum bicolor (L.) Moench], combining ability and heterosis analysis are commonly used to evaluate superior parental lines and to screen for strongly heterotic hybrids, which helps in sorghum variety selection and breeding. In this context, combining ability and heterosis analysis were assessed using 14 restorer lines and seven cytoplasmic male sterile (CMS) lines in 2019 and 2020. The analysis of variance of all cross combinations had highly significant differences for all characters studied, which indicated a wide variation across the parents, lines, testers, and crosses. Combining ability analysis showed that the general combining ability (GCA) and specific combining ability (SCA) of the different parents were differed significantly among different traits. Most combinations with high SCA also showed high GCA in their parent lines. The heritability in the narrow sense of grain weight per panicle and grain yield was relatively low, indicating that the ability of these traits to be directly inherited by offspring was weak, that they were greatly affected by the environment. The better-parent heterosis for plant height, grain weight per panicle, panicle length, and 1000-grain weight was consistent with the order of mid-parent heterosis from strong to weak. The GCA effects of two lines 10480A, 3765A and three testers 0-30R, R111, and JY15R were significant for the majority of the agronomic traits including grain yield and might be used for improving the yield of grains in sorghum as parents of excellent specific combining ability. Seven strongly heterotic F1 hybrids were screened; of these, hybrids 3765A × R111, 1102A × L2R, and 3765A × JY15R showed significant increases in seed iristectorigenin A content and will feature into the creation of new sorghum varieties rich in iristectorigenin A.

Effect of Renal Impairment on the Pharmacokinetics of a Single Oral Dose of Danuglipron in Participants With Type 2 Diabetes.

Danuglipron (PF-06882961) is an oral, small-molecule glucagon-like peptide-1 receptor agonist in development for the treatment of type 2 diabetes (T2D) and obesity. Impaired renal function is prevalent in patients with T2D. This Phase 1, open-label study evaluated the effect of renal impairment on the pharmacokinetics, safety, and tolerability of danuglipron (20 mg) in healthy participants with normal renal function (estimated glomerular filtration rate [eGFR] unnormalized for body surface area: ≥90 mL/min), in participants with T2D and normal renal function (eGFR ≥90 mL/min), and in participants with T2D and mild (eGFR 60-89 mL/min), moderate (eGFR 30-59 mL/min), or severe (eGFR <30 mL/min) renal impairment (N = 39). Log-linear regression analyses and analyses of variance showed no evidence of a clinically significant effect of reduced renal function on danuglipron pharmacokinetics. Renal clearance of unchanged danuglipron was minimal (<1% across all renal function groups). Danuglipron pharmacokinetics were similar between healthy participants and participants with T2D and normal renal function. A single 20-mg oral dose of danuglipron was generally safe and well tolerated in all participant groups. In participants with T2D, renal impairment had no clinically meaningful effect on the pharmacokinetic, safety, and tolerability profiles of danuglipron, indicating that dose adjustment of danuglipron will not be required when administered to patients with T2D and reduced renal function.

Microbes mediated immunogenic cell death in cancer immunotherapy.

Immunogenic cell death (ICD) is one of the 12 distinct cell death forms, which can trigger immune system to fight against cancer cells. During ICD, a number of cellular changes occur that can stimulate an immune response, including the release of molecules called damage-associated molecular patterns (DAMPs), signaling to immune cells to recognize and attack cancer cells. By virtue of their pivotal role in immune surveillance, ICD-based drug development has been a new approach to explore novel therapeutic combinations and personalized strategies in cancer therapy. Several small molecules and microbes can induce ICD-relevant signals and cause cancer cell death. In this review, we highlighted the role of microbe-mediate ICD in cancer immunotherapy and described the mechanisms through which microbes might serve as ICD inducers in cancer treatment. We also discussed current attempts to combine microbes with chemotherapy regimens or immune checkpoint inhibitors (ICIs) in the treatment of cancer patients. We surmise that manipulation of microbes may guide personalized therapeutic interventions to facilitate anticancer immune response.

Magnetic activated carbon from spent coffee grounds: iron-catalyzed CO2 activation mechanism and adsorption of antibiotic lomefloxacin from aqueous medium.

The facile fabrication of low-cost adsorbents possessing high removal efficiency and convenient separation property is an urgent need for water treatment. Herein, magnetic activated carbon was synthesized from spent coffee grounds (SCG) by Fe-catalyzed CO2 activation at 800 °C for 90 min, and magnetization and pore formation were simultaneously achieved during heat treatment. The sample was characterized by N2 adsorption-desorption, XRD, VSM, SEM, and FTIR. Batch adsorption experiments were conducted using lomefloxacin (LMO) as the probing pollutant. Preparation mechanism was revealed by TG-FTIR and XRD. Experimental results showed that Fe3O4 derived from Fe species can be reduced to Fe by carbon at high temperatures, followed by subsequent reoxidation to Fe3O4 by CO2, and the redox cycle between Fe and Fe3O4 favored the formation of pores. The promotion effects of Fe species on CO2 activation can be quantitatively reflected by the yield of CO as the signature gaseous product, and the suitable activation temperate range was determined to be 675 to 985 °C. The BET surface area, total pore volume, and saturated magnetization value of the product were 586 m2 g-1, 0.327 cm3 g-1, and 11.59 emu g-1, respectively. The Langmuir model was applicable for the adsorption isotherm data for LMO with the maximum adsorption capacity of 95 mg g-1, and thermodynamic analysis revealed that the adsorption process was endothermic and spontaneous. This study demonstrated that Fe-catalyzed CO2 activation was an effective method of converting SCG into magnetic separable adsorbent for LMO removal from aqueous medium.

Ultrasound-Assisted Enzymatic Protein Hydrolysis in Food Processing: Mechanism and Parameters.

Ultrasound has been widely used as a green and efficient non-thermal processing technique to assist with enzymatic hydrolysis. Compared with traditional enzymatic hydrolysis, ultrasonic-pretreatment-assisted enzymatic hydrolysis can significantly improve the efficiency of enzymatic hydrolysis and enhance the biological activity of substrates. At present, this technology is mainly used for the extraction of bioactive substances and the degradation of biological macromolecules. This review is focused on the mechanism of enzymatic hydrolysis assisted by ultrasonic pretreatment, including the effects of ultrasonic pretreatment on the enzyme structure, substrate structure, enzymatic hydrolysis kinetics, and thermodynamics and the effects of the ultrasonic conditions on the enzymatic hydrolysis results. The development status of ultrasonic devices and the application of ultrasonic-assisted enzymatic hydrolysis in the food industry are briefly described in this study. In the future, more attention should be paid to research on ultrasound-assisted enzymatic hydrolysis devices to promote the expansion of production and improve production efficiency.

Metabolism of glucose activates TORC1 through multiple mechanisms in Saccharomyces cerevisiae.

Target of Rapamycin Complex 1 (TORC1) is a conserved eukaryotic protein complex that links the presence of nutrients with cell growth. In Saccharomyces cerevisiae, TORC1 activity is positively regulated by the presence of amino acids and glucose in the medium. However, the mechanisms underlying nutrient-induced TORC1 activation remain poorly understood. By utilizing an in vivo TORC1 activation assay, we demonstrate that differential metabolism of glucose activates TORC1 through three distinct pathways in yeast. The first "canonical Rag guanosine triphosphatase (GTPase)-dependent pathway" requires conversion of glucose to fructose 1,6-bisphosphate, which activates TORC1 via the Rag GTPase heterodimer Gtr1GTP-Gtr2GDP. The second "non-canonical Rag GTPase-dependent pathway" requires conversion of glucose to glucose 6-phosphate, which activates TORC1 via a process that involves Gtr1GTP-Gtr2GTP and mitochondrial function. The third "Rag GTPase-independent pathway" requires complete glycolysis and vacuolar ATPase reassembly for TORC1 activation. We have established a roadmap to deconstruct the link between glucose metabolism and TORC1 activation.

Melatonin enhances salt tolerance in sorghum by modulating photosynthetic performance, osmoregulation, antioxidant defense, and ion homeostasis.

Melatonin is a potent antioxidant that can prevent plant damage caused by adverse stresses. It remains unclear whether exogenous melatonin can mitigate the effects of salt stress on seed germination and seedling growth of sorghum (Sorghum bicolor (L.) Moench). The aim of this study was to decipher the protective mechanisms of exogenous melatonin (100 µmol/L) on sorghum seedlings under NaCl-induced salt stress (120 mmol/L). Plant morphological, photosynthetic, and physiological characteristics were analyzed at different timepoints after sowing. Results showed that salt stress inhibited seed germination, seedling growth, and plant biomass accumulation by reducing photosynthetic pigment contents, photosynthetic efficiency, root vigor, and mineral uptake. In contrast, seed priming with melatonin enhanced photosynthetic pigment biosynthesis, photosynthetic efficiency, root vigor, and K+ content under salt stress. Melatonin application additionally enhanced the activities of antioxidant enzymes (superoxide dismutase, catalase, ascorbate peroxidase, and glutathione reductase) and increased the levels of non-enzymatic antioxidants (reduced glutathione, ascorbic acid) in the leaves. These changes were accompanied by increase in the leaf contents of soluble sugars, soluble proteins, and proline, as well as decrease in hydrogen peroxide accumulation, malondialdehyde content, and electrolyte leakage. Our findings indicate that exogenous melatonin can alleviate salt stress-induced damage in sorghum seedlings through multifaceted mechanisms, such as improving photosynthetic performance and root vigor, facilitating ion homeostasis and osmoregulation, and promoting antioxidant defense and reactive oxygen species scavenging.

Lateral single incision laparoscopic totally extraperitoneal hernioplasty (L-SILTEP) after laparoscopic radical prostatectomy: A rare case report with literature review.

INTRODUCTION: Single-incision laparoscopic totally extraperitoneal hernioplasty is a commonly used surgical procedure for the treatment of inguinal hernia. However, it is difficult to use traditional single incision laparoscopic totally extraperitoneal hernioplasty to treat inguinal hernia after laparoscopic radical prostatectomy. We successfully and smoothly cured a patient with left inguinal hernia after laparoscopic radical prostatectomy using lateral single incision laparoscopic totally extraperitoneal hernioplasty. CASE PRESENTATION: We report the case of a 70-year-old man who underwent laparoscopic radical prostatectomy 2 years earlier and had an evanescent mass in the left inguinal region for 1 month. DIAGNOSIS: On the basis of preoperative abdominal computed tomography and intraoperative findings, the patient was diagnosed with a left indirect inguinal hernia, and post-laparoscopic radical prostatectomy. INTERVENTIONS: The patient underwent lateral single incision laparoscopic totally extraperitoneal hernioplasty. OUTCOMES: The patient recovered well after the operation, and there were no postoperative complications or recurrence of inguinal hernia 3 months after the operation. CONCLUSION: For patients who have undergone laparoscopic radical prostatectomy, lateral single-incision laparoscopic totally extraperitoneal hernioplastycan be performed.

Sustainable utilization of Sedum plumbizincicola as superior hydrochar for efficient nutrients recovery.

To realize sound disposal of hyperaccumulator harvested from phytoremediation, hydrothermal carbonization (HTC) has been employed to obtain superior hydrochar adsorbents for removal of phosphate and ammonium from water body. A series of hydrochars have been prepared under tuned HTC conditions to tailor hydrochar with desired properties. Generally, increased temperature and prolonged reaction time facilitated acidic oxygen functional groups on hydrochars, thereby improving adsorption capacity of hydrochar. In single solute system, a superior hydrochar, derived from HTC under 260 °C for 2 h, achieved a maximum phosphate and ammonium adsorption capacity of 52.46 mg/g and 27.56 mg/g at 45 °C, respectively. In binary system, synergistic adsorption was observed only in lower solute concentration, whereas competitive adsorption occurred under higher solute concentration. Characterization and adsorption kinetics suggested chemisorption may dominate the adsorption process, thus the adsorption capacity could be improved by tuning pHpzc of hydrochar. This study firstly demonstrates the sustainable utilization of hyperaccumulators into nutrients-enriched hydrochar as fertilizer for in-situ phytoremediation of contaminated sites with minimized environmental risks towards circular economy.

Genome­wide identification and characterization of miR396 family members and their target genes GRF in sorghum (Sorghum bicolor (L.) moench).

MicroRNAs (miRNAs) widely participate in plant growth and development. The miR396 family, one of the most conserved miRNA families, remains poorly understood in sorghum. To reveal the evolution and expression pattern of Sbi-miR396 gene family in sorghum, bioinformatics analysis and target gene prediction were performed on the sequences of the Sbi-miR396 gene family members. The results showed that five Sbi-miR396 members, located on chromosomes 4, 6, and 10, were identified at the whole-genome level. The secondary structure analysis showed that the precursor sequences of all five Sbi-miR396 potentially form a stable secondary stem-loop structure, and the mature miRNA sequences were generated on the 5' arm of the precursors. Sequence analysis identified the mature sequences of the five sbi-miR396 genes were high identity, with differences only at the 1st, 9th and 21st bases at the 5' end. Phylogenetic analysis revealed that Sbi-miR396a, Sbi-miR396b, and Sbi-miR396c were clustered into Group I, and Sbi-miR396d and Sbi-miR396e were clustered into Group II, and all five sbi-miR396 genes were closely related to those of maize and foxtail millet. Expression analysis of different tissue found that Sbi-miR396d/e and Sbi-miR396a/b/c were preferentially and barely expressed, respectively, in leaves, flowers, and panicles. Target gene prediction indicates that the growth-regulating factor family members (SbiGRF1/2/3/4/5/6/7/8/10) were target genes of Sbi-miR396d/e. Thus, Sbi-miR396d/e may affect the growth and development of sorghum by targeting SbiGRFs. In addition, expression analysis of different tissues and developmental stages found that all Sbi-miR396 target genes, SbiGRFs, were barely expressed in leaves, root and shoot, but were predominantly expressed in inflorescence and seed development stage, especially SbiGRF1/5/8. Therefore, inhibition the expression of sbi-miR396d/e may increase the expression of SbiGRF1/5/8, thereby affecting floral organ and seed development in sorghum. These findings provide the basis for studying the expression of the Sbi-mir396 family members and the function of their target genes.

The Identification of a Yield-Related Gene Controlling Multiple Traits Using GWAS in Sorghum (Sorghum bicolor L.).

Sorghum bicolor (L.) is one of the oldest crops cultivated by human beings which has been used in food and wine making. To understand the genetic diversity of sorghum breeding resources and further guide molecular-marker-assisted breeding, six yield-related traits were analyzed for 214 sorghum germplasm from all over the world, and 2,811,016 single-nucleotide polymorphisms (SNPs) markers were produced by resequencing these germplasms. After controlling Q and K, QTLs were found to be related to the traits using three algorisms. Interestingly, an important QTL was found which may affect multiple traits in this study. It was the most likely candidate gene for the gene SORBI_3008G116500, which was a homolog of Arabidopsis thaliana gene-VIP5 found by analyzing the annotation of the gene in the LD block. The haplotype analysis showed that the SORBI_3008G116500hap3 was the elite haplotype, and it only existed in Chinese germplasms. The traits were proven to be more associated with the SNPs of the SORBI_3008G116500 promoter through gene association studies. Overall, the QTLs and the genes identified in this study would benefit molecular-assisted yield breeding in sorghum.

Modern water at low latitudes on Mars: Potential evidence from dune surfaces.

Landforms on the Martian surface are critical to understanding the nature of surface processes in the recent past. However, modern hydroclimatic conditions on Mars remain enigmatic, as explanations for the formation of observed landforms are ambiguous. We report crusts, cracks, aggregates, and bright polygonal ridges on the surfaces of hydrated salt-rich dunes of southern Utopia Planitia (~25°N) from in situ exploration by the Zhurong rover. These surface features were inferred to form after 1.4 to 0.4 million years ago. Wind and CO2 frost processes can be ruled out as potential mechanisms. Instead, involvement of saline water from thawed frost/snow is the most likely cause. This discovery sheds light on more humid conditions of the modern Martian climate and provides critical clues to future exploration missions searching for signs of extant life, particularly at low latitudes with comparatively warmer, more amenable surface temperatures.

ß-Elemene enhances erlotinib sensitivity through induction of ferroptosis by upregulating lncRNA H19 in EGFR-mutant non-small cell lung cancer.

Nearly half of all Asian non-small cell lung cancer (NSCLC) patients harbour epidermal growth factor receptor (EGFR) mutations, and first-generation EGFR tyrosine kinase inhibitors (TKIs) are one of the first-line treatments that have improved the outcomes of these patients. Unfortunately, 20% of these patients can not benefit from the treatment. The basis of this primary resistance is poorly understood. Therefore, overcoming EGFR-TKI primary resistance and maintaining the efficacy of TKIs has become a key issue. ß-Elemene, a sesquiterpene compound extracted from Curcuma aromatica Salisb. (wenyujing), has shown potent antitumor effects. In this research, we found that ß-elemene combined with erlotinib enhanced the cytotoxicity of erlotinib to primary EGFR-TKI-resistant NSCLC cells with EGFR mutations and that ferroptosis was involved in the antitumor effect of the combination treatment. We found that lncRNA H19 was significantly downregulated in primary EGFR-TKI-resistant NSCLC cell lines and was upregulated by the combination treatment. Overexpression or knockdown of H19 conferred sensitivity or resistance to erlotinib, respectively, in both in vitro and in vivo studies. The high level of H19 enhanced the cytotoxicity of erlotinib by inducing ferroptosis. In conclusion, our data showed that ß-elemene combined with erlotinib could enhance sensitivity to EGFR-TKIs through induction of ferroptosis via H19 in primary EGFR-TKI-resistant lung cancer, providing a promising strategy to overcome EGFR-TKI resistance in NSCLC patients.

Modified Flowing Material Balance Equation for Shale Gas Reservoirs.

To determine original gas-in-place, this study establishes a flowing material balance equation based on the improved material balance equation for shale gas reservoirs. The method considers the free gas in the matrix and fracture, the dissolved gas in kerogen, and the pore volume occupied by adsorbed phase simultaneously, overcoming the problem of incomplete consideration in the earlier models. It also integrates the material balance method with the flowing material balance method to obtain the average formation pressure, eliminating the problem with the previous method where shutting down of wells was needed to monitor the formation pressure. The volume of the adsorbed gas on the ground is converted into volume of the adsorbed phase in the formation using the volume conservation method to characterize the pore volume occupied by the adsorbed phase, which solves the problem of the previous model that the adsorbed phase was neglected in the pore volume. The model proposed in this study is applied to the Fuling Shale Gas Field in southwest China and compared with other flowing material balance equations, and the results show that the single-well control area calculated by the model proposed in this study is closer to the real value, indicating that the calculations in this study are more accurate. Furthermore, the calculations show that the dissolved gas takes up a large fraction of the total reserves and cannot be ignored. The sensitivity analyses of critical parameters demonstrate that (a) the greater the porosity of the fracture, the greater the free gas storage; (b) the values of Langmuir volume and TOC can significantly affect the results of the reservoir calculation; and (c) the adsorbed phase occupies a smaller pore volume when the Langmuir volume is smaller, the Langmuir pressure is higher, or the adsorbed phase density is higher. The findings of this study can provide better understanding of the necessity to take into account the dissolved gas in the kerogen, the pore volume occupied by the adsorbed phase, and the fracture porosity when evaluating reserves. The method could be applied to the calculation of pressure, recovery of free gas phase and adsorbed phase, original gas-in-place, and production predictions, which could help for better guidance of reserve potential estimations and development strategies of shale gas reservoirs.