Growth arrest DNA damage-inducible protein 45 β (GADD45B) has been reported to be a regulatory factor for active DNA demethylation and is implicated in the modulation of synaptic plasticity and chronic stress-related psychopathological processes. However, its precise role and mechanism of action in stress susceptibility remain elusive. In this study, we found a significant reduction in GADD45B expression specifically in the ventral, but not the dorsal hippocampal CA1 (dCA1) of stress-susceptible mice. Furthermore, we demonstrated that GADD45B negatively regulates susceptibility to social stress and NMDA receptor-dependent long-term potentiation (LTP) in the ventral hippocampal CA1 (vCA1). Importantly, through pharmacological inhibition using the NMDA receptor antagonist MK801, we provided further evidence supporting the hypothesis that GADD45B potentially modulates susceptibility to social stress by influencing NMDA receptor-mediated LTP. Collectively, these results suggested that modulation of NMDA receptor-mediated synaptic plasticity is a pivotal mechanism underlying the regulation of susceptibility to social stress by GADD45B.
Animals ; Receptors, N-Methyl-D-Aspartate/antagonists & inhibitors* ; CA1 Region, Hippocampal/drug effects* ; Male ; Stress, Psychological/physiopathology* ; Mice ; Neuronal Plasticity/drug effects* ; Long-Term Potentiation/drug effects* ; Mice, Inbred C57BL ; Antigens, Differentiation/metabolism* ; Dizocilpine Maleate/pharmacology* ; Excitatory Amino Acid Antagonists/pharmacology* ; GADD45 Proteins

Chronic visceral pain is a persistent and debilitating condition arising from dysfunction or sensitization of the visceral organs and their associated nervous pathways. Increasing evidence suggests that imbalances in central nervous system function play an essential role in the progression of visceral pain, but the exact mechanisms underlying the neural circuitry and molecular targets remain largely unexplored. In the present study, the ventral tegmental area (VTA) was shown to mediate visceral pain in mice. Visceral pain stimulation increased c-Fos expression and Ca²⁺ activity of glutamatergic VTA neurons, and optogenetic modulation of glutamatergic VTA neurons altered visceral pain. In particular, the upregulation of NMDA receptor 2A (NR2A) subunits within the VTA resulted in visceral pain in mice. Administration of a selective NR2A inhibitor decreased the number of visceral pain-induced c-Fos positive neurons and attenuated visceral pain. Pharmacology combined with chemogenetics further demonstrated that glutamatergic VTA neurons regulated visceral pain behaviors based on NR2A. In summary, our findings demonstrated that the upregulation of NR2A in glutamatergic VTA neurons plays a critical role in visceral pain. These insights provide a foundation for further comprehension of the neural circuits and molecular targets involved in chronic visceral pain and may pave the way for targeted therapies in chronic visceral pain.
Animals ; Male ; Visceral Pain/metabolism* ; Up-Regulation/physiology* ; Ventral Tegmental Area/metabolism* ; Receptors, N-Methyl-D-Aspartate/antagonists & inhibitors* ; Neurons/drug effects* ; Mice, Inbred C57BL ; Mice ; Proto-Oncogene Proteins c-fos/metabolism* ; Chronic Pain/metabolism* ; Glutamic Acid/metabolism*

Empathy is crucial for communication and survival for individuals. Whether empathy in pain contagion shows sex differences and its underlying mechanisms remain unclear. Here, we report that pain contagion can occur in stranger female rats, but not in stranger males. Blocking oxytocin receptors in the anterior cingulate cortex (ACC) suppressed pain contagion in female strangers, while oxytocin administration induced pain contagion in male strangers. In vitro, corticosterone reduces neuronal activation by oxytocin. During male stranger interactions, higher corticosterone decreased oxytocin receptor-positive neuronal activity in the ACC, suppressing pain contagion. These findings highlight the role of oxytocin in pain contagion and suggest that sex differences in empathy may be determined by the balance of oxytocin and corticosterone in the ACC. This study suggests an approach for the treatment of certain mental disorders associated with abnormal empathy, such as autism and depression.
Animals ; Oxytocin/pharmacology* ; Gyrus Cinguli/drug effects* ; Male ; Female ; Corticosterone/pharmacology* ; Empathy/drug effects* ; Sex Characteristics ; Receptors, Oxytocin/antagonists & inhibitors* ; Pain/psychology* ; Rats ; Rats, Sprague-Dawley ; Neurons/metabolism*

Hypoxia and aberrant activation of epidermal growth factor receptor (EGFR) are considered important features of various malignancies. However, whether hypoxia can directly trigger EGFR activation and its clinical implications remain unclear. In this study, we demonstrated that in oral cancer, a typical hypoxic tumor, hypoxia can induce chronic but constitutive phosphorylation of wild-type EGFR in the absence of ligands. Oral cancer cell lines exhibit different EGFR phosphorylation responses to hypoxia. In hypoxic HN4 and HN6 cells, ubiquitination-mediated endocytosis, lysosomal sorting, and degradation lead to low levels of EGFR phosphorylation. However, in CAL-27 and HN30 cells, a novel HIF-1α-induced long noncoding RNA (lncRNA), EUDAL, can compete with the E3 ligase/adaptor complex c-Cbl/Grb2 for binding to EGFR, stabilizing phosphorylated EGFR (pEGFR) and resulting in sustained activation of EGFR and its downstream STAT3/BNIP3 signaling. STAT3/BNIP3-mediated autophagy leads to antitumor drug resistance. A high EUDAL/EGFR/STAT3/autophagy pathway activation predicts poor response to chemotherapy in oral cancer patients. Collectively, hypoxia can induce noncanonical ligand-independent EGFR phosphorylation. High EUDAL expression facilitates sustained EGFR phosphorylation in hypoxic tumor cells and leads to autophagy-related drug resistance.
Humans ; ErbB Receptors/metabolism* ; Mouth Neoplasms/pathology* ; RNA, Long Noncoding/genetics* ; Drug Resistance, Neoplasm/genetics* ; Cell Line, Tumor ; Phosphorylation ; Signal Transduction ; STAT3 Transcription Factor/metabolism* ; Cell Hypoxia ; Autophagy ; Proto-Oncogene Proteins c-cbl/metabolism*

Polycystic ovary syndrome (PCOS) is the predominant cause of subfertility in reproductive-aged women; however, its pathophysiology remains unknown. Neurotensin (NTS) is a member of the gut-brain peptide family and is involved in ovulation; its relationship with PCOS is unclear. Here, we found that NTS expression in ovarian granulosa cells and follicular fluids was markedly decreased in patients with PCOS. In the in vitro culture of cumulus-oocyte complexes, the neurotensin receptor 1 (NTSR1) antagonist SR48692 blocked cumulus expansion and oocyte meiotic maturation by inhibiting metabolic cooperation and damaging the mitochondrial structure in oocytes and surrounding cumulus cells. Furthermore, the ERK1/2-early growth response 1 pathway was found to be a key downstream mediator of NTS/NTSR1 in the ovulatory process. Animal studies showed that in vivo injection of SR48692 in mice reduced ovulation efficiency and contributed to irregular estrus cycles and polycystic ovary morphology. By contrast, NTS partially ameliorated the ovarian abnormalities in mice with dehydroepiandrosterone-induced PCOS. Our findings highlighted the critical role of NTS reduction and consequent abnormal NTSR1 signaling in the ovulatory dysfunction of PCOS, suggesting a potential strategy for PCOS treatment.
Polycystic Ovary Syndrome/physiopathology* ; Female ; Animals ; Neurotensin/metabolism* ; Receptors, Neurotensin/antagonists & inhibitors* ; Mice ; Ovulation/drug effects* ; Humans ; Granulosa Cells/metabolism* ; Adult ; Oocytes/metabolism* ; MAP Kinase Signaling System ; Signal Transduction ; Follicular Fluid/metabolism* ; Disease Models, Animal ; Gonadotropin-Releasing Hormone/analogs & derivatives*

To investigate the protective effects and underlying mechanisms of Qin-Zhu-Liang-Xue decoction (QZLX) in atopic dermatitis (AD) and glucocorticoid resistance, we conducted a single-blinded, randomized controlled clinical trial to evaluate the efficacy and safety of this concoction. Network pharmacology analysis was performed and validated through clinical studies. The efficacy, safety, and mechanism of action of QZLX and glucocorticoid receptor (GR) α recombinant protein were assessed in AD mice induced by 2,4-dinitrofluorobenzene (DNFB). Correlation analysis was performed to determine the clinical relevance of GRα. The trial demonstrated that patients who received QZLX showed considerable improvements in their Scoring Atopic Dermatitis (SCORAD) and Dermatology Life Quality Index (DLQI) scores compared with those who received mizolastine at week 4. Network pharmacological analysis identified GRα as a key target for QZLX in AD treatment. QZLX administration increased the serum GRα expression in AD patients, alleviated AD symptoms in mice, decreased inflammatory cytokine expression, and increased GRα expression without affecting liver or kidney function. In addition, GRα recombinant protein improved AD-like skin lesions in DNFB-induced mice. A negative correlation was observed between GRα expression and clinical parameters, including SCORAD, DLQI, and serum IgE levels. QZLX alleviates AD symptoms through the upregulation of GRα and thus presents a novel therapeutic strategy for the prevention of glucocorticoid resistance in AD management.
Dermatitis, Atopic/drug therapy* ; Animals ; Drugs, Chinese Herbal/administration & dosage* ; Humans ; Mice ; Network Pharmacology ; Male ; Female ; Adult ; Receptors, Glucocorticoid/metabolism* ; Disease Models, Animal ; Single-Blind Method ; Middle Aged ; Young Adult

Therapeutic resistance in cancer is responsible for numerous cancer deaths in clinical practice. While target mutations are well recognized as the basis of genetic resistance to targeted therapy, nontarget mutation resistance (or nongenetic resistance) remains poorly characterized. Despite its complex and unintegrated mechanisms in the literature, nongenetic resistance is considered from our perspective to be a collective response of innate or acquired resistant subpopulations in heterogeneous tumors to therapy. These subpopulations, e.g., cancer stem-like cells, cancer cells with epithelial-to-mesenchymal transition, and drug-tolerant persisters, are protected by their resistance traits at cellular and molecular levels. This review summarizes recent advances in the research on resistant populations and their resistance traits. NOTCH signaling, as a central regulator of nongenetic resistance, is discussed with a special focus on its canonical maintenance of resistant cancer cells and noncanonical regulation of their resistance traits. This novel view of canonical and noncanonical NOTCH signaling pathways is translated into our proposal of reshaping therapeutic strategies targeting NOTCH signaling in resistant cancer cells. We hope that this review will lead researchers to study the canonical and noncanonical arms of NOTCH signaling as an integrated resistant mechanism, thus promoting the development of innovative therapeutic strategies.
Neoplasms/metabolism* ; Receptors, Notch/metabolism* ; Disease Resistance/physiology* ; Signal Transduction/physiology* ; Humans ; Drug Resistance, Neoplasm/physiology* ; Molecular Targeted Therapy/methods*

Based on the epidermal growth factor receptor(EGFR)/mitogen-activated protein kinase(MAPK) signaling pathway-mediated cell proliferation, this study explores the effect of cisplatin combined with Guiqi Yiyuan Ointment on Lewis lung cancer-bearing mice. A total of 60 male C57BL/6 mice were randomly divided into a blank group with 10 mice and a modeling group with 50 mice. After modeling, they were randomly divided into the model group, cisplatin group, and low-, medium-, and high-dose groups of cisplatin combined with Guiqi Yiyuan Ointment, with 10 mice in each group. After 14 days of medication, the general condition of the mice was observed; body weight was measured, and organ index and tumor inhibition rate were calculated. Hematoxylin-eosin(HE) staining was used to observe the pathological morphology changes in tumor tissue. Immunohistochemistry was used to detect the positive rate of Ki-67 antigen(Ki-67) and proliferating cell nuclear antigen(PCNA) in tumor tissue. Western blot and real time-quantitative polymerase chain reaction(qPCR) were used to detect the expression of related proteins and mRNA in tumor tissue. Flow cytometry was used to detect the cell cycle of tumor cells in tumor tissue. The results showed that compared with that in the blank group, the general condition of mice in the model group deteriorated; the body weight, as well as thymus and spleen index of mice in the model group decreased after 14 days of medication. Compared with that in the model group, the general condition of mice in the cisplatin group deteriorated, while the condition of mice in the combined groups improved; the body weight, as well as thymus and spleen index of mice in the cisplatin group decreased, while the three indicators in the combined groups increased; the tumor weight of each medication group decreased, and the tumor inhibition rate increased; there were varying degrees of necrosis in tumor cells of each medication group, and the tightness of tumor cells, the increase in the number of cell nuclei and chromatin, and mitosis all decreased. The positive rate of Ki-67 and PCNA, as well as the protein expression and ratio of p-EGFR/EGFR, rat sarcoma viral oncogene homolog(Ras), phosphorylated Raf-1 protein kinase(p-Raf-1)/Raf-1, phosphorylated mitogen-activated protein kinase kinase(p-MEK)/MEK, phosphorylated extracellular signal-regulated kinase(p-ERK)/ERK and the mRNA expression of EGFR, Ras, Raf-1, MEK, and ERK all decreased. The proportion of tumor cells in the G_0/G_1 phase of each medication group increased, and that in the S phase decreased. In addition, there was no significant difference in the G_2/M phase. Compared with that of the cisplatin group, the tumor weight of the combined groups decreased, and the tumor inhibition rate increased. The necrosis and mitosis of tumor cells in the combined groups were more pronounced; the positive rate of Ki-67 and PCNA, the protein expression and ratio of p-EGFR/EGFR, Ras, p-Raf-1/Raf-1, p-MEK/MEK, and p-ERK/ERK, as well as the mRNA expression of EGFR, Ras, Raf-1, MEK, and ERK in the combined groups all decreased. The proportion of tumor cells in the G_0/G_1 phase of the combined medium-and high-dose groups increased, and that in the S phase decreased. There was no significant difference in the proportion of tumor cells of the combined groups in the G_2/M phase. This indicates that the combination of cisplatin and Guiqi Yiyuan Ointment can enhance the anti-tumor effect of cisplatin on tumor-bearing mice, and the mechanism may be associated with the inhibition of the EGFR/MAPK pathway, which accelerates the arrest of tumor cells in the G_0/G_1 phase, thereby inhibiting the proliferation of tumor cells. At the same time, the study also indicates that Guiqi Yiyuan Ointment may reduce the damage of tumors to mice and the toxic side effects brought by cisplatin chemotherapy.
Animals ; Male ; Carcinoma, Lewis Lung/metabolism* ; Drugs, Chinese Herbal/administration & dosage* ; ErbB Receptors/genetics* ; Mice ; Cisplatin/administration & dosage* ; Mice, Inbred C57BL ; Cell Proliferation/drug effects* ; Ointments/administration & dosage* ; MAP Kinase Signaling System/drug effects* ; Humans ; Antineoplastic Agents/administration & dosage* ; Lung Neoplasms/metabolism*

In order to investigate whether the effect of Yuxuebi Tablets on the peripheral and central inflammation resolution of mice with inflammatory pain is related to their regulation of G protein-coupled receptor 37(GPR37), an inflammatory pain model was established by injecting complete Freund's adjuvant(CFA) into the paws of mice, with a sham-operated group receiving a similar volume of normal saline. The mice were assigned randomly to the sham-operated group, model group, ibuprofen group(91 mg·kg~(-1)), and low-, medium-, and high-dose groups of Yuxuebi Tablets(60, 120, and 240 mg·kg~(-1)). The drug was administered orally from days 1 to 19 after modeling. Von Frey method and the hot plate test were used to detect mechanical pain thresholds and heat hyperalgesia. The levels of interleukin-10(IL-10) and transforming growth factor-beta(TGF-β) in the spinal cord were quantified using enzyme-linked immunosorbent assay(ELISA), and the mRNA and protein expression of GPR37 in the spinal cord was measured by real-time quantitative reverse transcription PCR(qRT-PCR) and Western blot. Additionally, immunofluorescence was used to detect the expression of macrosialin antigen(CD68), mannose receptor(MRC1 or CD206), and GPR37 in dorsal root ganglia, as well as the expression of calcium-binding adapter molecule 1(IBA1), CD206, and GPR37 in the dorsal horn of the spinal cord. The results showed that compared with those of the sham-operated group, the mechanical pain thresholds and hot withdrawal latency of the model group significantly declined, and the expression of CD68 in the dorsal root ganglia and the expression of IBA1 in the dorsal horn of the spinal cord significantly increased. The expression of CD206 and GPR37 significantly decreased in the dorsal root ganglion and dorsal horn of the spinal cord, and IL-10 and TGF-β levels in the spinal cord were significantly decreased. Compared with those of the model group, the mechanical pain thresholds and hot withdrawal latency of the high-dose group of Yuxuebi Tablets significantly increased, and the expression of CD68 in the dorsal root ganglion and IBA1 in the dorsal horn of the spinal cord significantly decreased. The expression of CD206 and GPR37 in the dorsal root ganglion and dorsal horn of the spinal cord significantly increased, as well as IL-10 and TGF-β levels in the spinal cord. These findings indicated that Yuxuebi Tablets may reduce macrophage(microglial) infiltration and foster M2 macrophage polarization by enhancing GPR37 expression in the dorsal root ganglia and dorsal horn of the spinal cord of CFA-induced mice, so as to improve IL-10 and TGF-β levels, promote resolution of both peripheral and central inflammation, and play analgesic effects.
Inflammation/genetics* ; Pain/genetics* ; Drugs, Chinese Herbal/administration & dosage* ; Animals ; Mice ; Freund's Adjuvant/pharmacology* ; Ibuprofen ; Pain Threshold/drug effects* ; Hyperalgesia/genetics* ; Ganglia, Spinal ; Interleukin-10/genetics* ; Transforming Growth Factor beta/genetics* ; Reverse Transcriptase Polymerase Chain Reaction ; Tablets ; Receptors, G-Protein-Coupled

This study aims to investigate the potential effect of the water extract of fresh Rehmanniae Radix on hypercholesterolemia in mice that was induced by a high-fat and high-cholesterol diet and explore its possible mechanism from bile acid reabsorption. Male C57BL/6 mice were randomly assigned into the following groups: control, model, low-and high-dose(4 and 8 g·kg~(-1), respectively) fresh Rehmanniae Radix, and positive drug(simvastatin, 0.05 g·kg~(-1)). Other groups except the control group were fed with a high-fat and high-cholesterol diet for 6 consecutive weeks to induce hypercholesterolemia. From the 6th week, mice were administrated with corresponding drugs daily via gavage for additional 6 weeks, while continuing to be fed with a high-fat and high-cholesterol diet. Serum levels of total cholesterol(TC), triglycerides(TG), low density lipoprotein-cholesterol(LDL-c), high density lipoprotein-cholesterol(HDL-c), and total bile acid(TBA), as well as liver TC and TG levels and fecal TBA level, were determined by commercial assay kits. Hematoxylin-eosin(HE) staining, oil red O staining, and transmission electron microscopy were performed to observe the pathological changes in the liver. Three livers samples were randomly selected from each of the control, model, and high-dose fresh Rehmanniae Radix groups for high-throughput transcriptome sequencing. Differentially expressed genes were mined and KEGG pathway enrichment analysis was performed to predict the key pathways and target genes of the water extract of fresh Rehmanniae Radix in the treatment of hypercholesterolemia. RT-qPCR was employed to measure the mRNA levels of cholesterol 7α-hydroxylase(CYP7A1) and cholesterol 27α-hydroxylase(CYP27A1) in the liver. Western blot was employed to determine the protein levels of CYP7A1 and CYP27A1 in the liver as well as farnesoid X receptor(FXR), apical sodium-dependent bile acid transporter(ASBT), and ileum bile acid-binding protein(I-BABP) in the ileum. The results showed that the water extract of fresh Rehmanniae Radix significantly lowered the levels of TC and TG in the serum and liver, as well as the level of LDL-c in the serum. Conversely, it elevated the level of HDL-c in the serum and TBA in feces. No significant difference was observed in the level of TBA in the serum among groups. HE staining, oil red O staining, and transmission electron microscopy showed that the water extract reduced the accumulation of lipid droplets in the liver. Further mechanism studies revealed that the water extract of fresh Rehmanniae Radix significantly down-regulated the protein levels of FXR and bile acid reabsorption-related proteins ASBT and I-BABP. Additionally, it enhanced CYP7A1 and CYP27A1, the key enzymes involved in bile acid synthesis. Therefore, it is hypothesized that the water extract of fresh Rehmanniae Radix may exert an anti-hypercholesterolemic effect by regulating FXR/ASBT/I-BABP signaling, inhibiting bile acid reabsorption, and increasing bile acid excretion, thus facilitating the conversion of cholesterol to bile acids.
Animals ; Male ; Bile Acids and Salts/metabolism* ; Mice, Inbred C57BL ; Mice ; Diet, High-Fat/adverse effects* ; Cholesterol/metabolism* ; Drugs, Chinese Herbal/administration & dosage* ; Hypercholesterolemia/genetics* ; Receptors, Cytoplasmic and Nuclear/genetics* ; Rehmannia/chemistry* ; Liver/drug effects* ; Humans ; Cholesterol 7-alpha-Hydroxylase/genetics* ; Plant Extracts