Central nervous system (CNS) World Health Organization (WHO) grade 3 meningiomas are aggressive tumors of the CNS whose clinical management is particularly challenging due to the lack of effective systemic treatment options. Recent multiomic studies revealed increasing expression and activity of the histone methyltransferase enhancer of zeste homolog 2 (EZH2) in parallel with the increase in meningioma grade. This study primarily aims to characterize in detail the clinical and molecular correlates of EZH2 expression in grade 3 meningiomas and to investigate its functional role both in vitro and in vivo. EZH2 expression was correlated with clinical outcome data as well as with the loss of p16 and H3 K27me3, 9p21.3 deletion, and pTERT mutational status in a cohort of 49 grade 3 meningiomas. Effects of the EZH2 methyltransferase inhibitor EPZ-6438 were investigated on the IOMM-Lee human grade 3 meningioma cell line. Higher levels of EZH2 overexpression were associated with shorter overall and local progression-free survival and higher proliferative activity in our grade 3 meningioma cohort. Deletion of the 9p21.3 region and loss of H3 K27me3 were related to higher EZH2 expression, while loss of p16 and pTERT mutations did not show significant correlation. EPZ-6438 treatment exerted concentration-dependent inhibitory effect on IOMM-Lee cell proliferation through inhibition of cell cycle progression, increased p21, and decreased FOXM1 expression. Upon per os EPZ-6438 treatment, a slight inhibition of tumor growth with different underlying mechanisms was detected in a mouse heterotopic xenograft model. Our findings demonstrate that EZH2 is functionally relevant in grade 3 meningiomas, primarily through promoting cell proliferation.

Bone cancer and its related chronic pain are huge clinical problems since the available drugs are often ineffective or cannot be used long term due to a broad range of side effects. The mechanisms, mediators and targets need to be identified to determine potential novel therapies. Here, we characterize a mouse bone cancer model induced by intratibial injection of K7M2 osteosarcoma cells using an integrative approach and investigate the role of capsaicin-sensitive peptidergic sensory nerves. The mechanical pain threshold was assessed by dynamic plantar aesthesiometry, limb loading by dynamic weight bearing, spontaneous pain-related behaviors via observation, knee diameter with a digital caliper, and structural changes by micro-CT and glia cell activation by immunohistochemistry in BALB/c mice of both sexes. Capsaicin-sensitive peptidergic sensory neurons were defunctionalized by systemic pretreatment with a high dose of the transient receptor potential vanilloid 1 (TRPV1) agonist resiniferatoxin (RTX). During the 14- and 28-day experiments, weight bearing on the affected limb and the paw mechanonociceptive thresholds significantly decreased, demonstrating secondary mechanical hyperalgesia. Signs of spontaneous pain and osteoplastic bone remodeling were detected both in male and female mice without any sex differences. Microglia activation was shown by the increased ionized calcium-binding adapter molecule 1 (Iba1) immunopositivity on day 14 and astrocyte activation by the enhanced glial fibrillary acidic protein (GFAP)-positive cell density on day 28 in the ipsilateral spinal dorsal horn. Interestingly, defunctionalization of the capsaicin-sensitive afferents representing approximately 2/3 of the nociceptive fibers did not alter any functional parameters. Here, we provide the first complex functional and morphological characterization of the K7M2 mouse osteosarcoma model. Bone-cancer-related chronic pain and hyperalgesia are likely to be mediated by central sensitization involving neuroinflammation via glial cell activation in the spinal dorsal horn, but not the capsaicin-sensitive sensory neuronal system.

Succinate dehydrogenase (SDH)-deficient renal cell carcinoma (RCC) is a rare subtype of renal neoplasm predominantly affecting younger individuals. It is characterized by germline mutations in SDHx genes, particularly type B. Histologically, SDH-deficient RCC features eosinophilic cytoplasmic cells forming solid nests or microcysts, sometimes entrapping normal tubules. We present three SDH-deficient RCC cases with overlapping morphological features with fumarate hydratase-deficient RCC and TFEB-rearranged RCC, an appearance that has not been previously described. All tumors lacked SDHB expression and harbored pathogenic SDHB mutations, with the germline nature confirmed in two cases. Metastasis developed in two patients. Our case set highlights the diagnostic challenges of molecularly defined renal tumors and expands the morphological spectrum of SDH-deficient RCC with unusual histological features. Clinically, these tumors appear to be aggressive.

Lysophosphatidic acid (LPA) is a bioactive phospholipid mediator that has been found to ameliorate nonsteroidal anti-inflammatory drug (NSAID)-induced gastric injury by acting on lysophosphatidic acid type 2 receptor (LPAR2). In this study, we investigated whether LPAR2 signaling was implicated in the development of NSAID-induced small intestinal injury (enteropathy), another major complication of NSAID use. Wild-type (WT) and Lpar2 deficient ( Lpar2 −/− ) mice were treated with a single, large dose (20 or 30 mg/kg, i.g.) of indomethacin (IND). The mice were euthanized at 6 or 24 h after IND treatment. We showed that IND-induced mucosal enteropathy and neutrophil recruitment occurred much earlier (at 6 h after IND treatment) in Lpar2 −/− mice compared to WT mice, but the tissue levels of inflammatory mediators (IL-1β, TNF-α, inducible COX-2, CAMP) remained at much lower levels. Administration of a selective LPAR2 agonist DBIBB (1, 10 mg/kg, i.g., twice at 24 h and 30 min before IND treatment) dose-dependently reduced mucosal injury and neutrophil activation in enteropathy, but it also enhanced IND-induced elevation of several proinflammatory chemokines and cytokines. By assessing caspase-3 activation, we found significantly increased intestinal apoptosis in IND-treated Lpar2 −/− mice, but it was attenuated after DBIBB administration, especially in non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice. Finally, we showed that IND treatment reduced the plasma activity and expression of autotaxin (ATX), the main LPA-producing enzyme, and also reduced the intestinal expression of Lpar2 mRNA, which preceded the development of mucosal damage. We conclude that LPAR2 has a dual role in NSAID enteropathy, as it contributes to the maintenance of mucosal integrity after NSAID exposure, but also orchestrates the inflammatory responses associated with ulceration. Our study suggests that IND-induced inhibition of the ATX-LPAR2 axis is an early event in the pathogenesis of enteropathy.

INTRODUCTION:Recent advances in the subclassification of small cell lung carcinomas (SCLCs) may help to overcome the unmet need for targeted therapies and improve survival. However, limited information is available on how the expression of the subtype markers changes during tumour progression. Our study aimed to compare the expression of these markers in primary and brain metastatic SCLCs. MATERIALS AND METHODS:Immunohistochemical analysis of the subtype markers was performed on 120 SCLCs (including 10 matched samples) and SCLC xenografts. RESULTS:Compared to primary SCLCs, there was a significant increase in the proportion of mixed subtypes in brain metastases, with a rate of ASCL1high/NeuroD1high and ASCL1high/NeuroD1high/YAP1high subtypes increasing to 48 % and 18 %, respectively. The subtype of the paired samples matched in only one-third of the cases. Although we did not observe a significant change after chemotherapy, a continuous decrease in ASCL1 expression coupled with an increase in the NeuroD1 expression was detected in the xenografts in a long-term experiment. DISCUSSION:Our results indicate that the expression of subtype markers frequently changes during disease progression, and subtype analysis of the primary SCLC may not provide accurate information about the characteristics of the recurrent or metastatic tumour. Therefore, repeated sampling and subtyping may be necessary for subtype-specific targeted therapy.

Glioblastomas are the most common IDH-wildtype adult high-grade gliomas, frequently harboring mutations in the promoter region of the TERT gene (pTERT) and utilizing the subsequent telomerase overexpression for telomere length maintenance. However, some rare cases show loss of ATRX and use alternative lengthening of telomeres. We aimed to perform the first complex genetic analysis specifically concentrating on the latter subgroup, since the molecular properties including potential clinically relevant features are poorly characterized. Comprehensive genomic profiling (CGP) of 12 ATRX-deficient and 13 ATRX-intact IDH-wildtype adult high-grade gliomas was performed using the Illumina TruSight Oncology 500 targeted next-generation sequencing panel. ATRX and pTERT mutations were revealed to be mutually exclusive by CGP. DNMT3A alterations were confined to ATRX-deficient, while PTEN mutations to ATRX-intact cases. EGFR amplification was relatively rare, while alterations of the RAS-MAPK pathway, including NF1 mutations and BRAF alterations, were more characteristic in the ATRX-deficient group. Several pathogenic or likely pathogenic variants of genes related to homologous recombination repair were detected in both groups, but with different patterns of affected genes. Two ATRX-deficient tumors with high tumor mutational burden and mismatch repair deficiency were found. One of these showed a peculiar association of oligodendroglioma-like morphology, novel fusions involving the NTRK2 and LRRFIP2 genes, POLE mutations as well as therapy-induced MLH1 and PMS2 loss. The other showed loss of MSH2 and MSH6 without genetic alterations in the encoding genes suggesting an epigenetic background. Apparent, statistically significant differences in the genetic characteristics of ATRX-deficient and ATRX-intact IDH-wildtype adult high-grade gliomas were revealed by our study. These observations suggest that tumors from the former group are particularly intriguing targets of potential future therapeutic interventions including immunotherapies combined with MAPK pathway inhibition and DNA-repair inhibitors.

Considerable changes were introduced into the 5th World Health Organization (WHO) classification of central nervous system (CNS) tumors, published in 2021, including new entities, a clearer classification of previous categories, correlating better with clinical behavior and changes in nomenclature. The number of definitions based on molecular features in addition to histopathology continued to increase. Here, we highlight the basic principles of the 5th CNS WHO classification and discuss glial, glioneuronal, neuronal, choroid plexus, embryonal and pineal tumors, as well as meningiomas in more details. We pay special attention to new entities as well as altered criteria and designations. Our primary goal is to present the "classical" pathological aspects, but the inseparable molecular pathological features are also briefly discussed, to the absolutely necessary extent for comprehension. We aim to provide a guideline to understand the modern classification of CNS tumors for practitioners of neuro-oncology and neuropathology.

The Tac4 gene-derived hemokinin-1 (HK-1) binds to the NK1 receptor, similarly to Substance P, and plays a role in acute stress reactions and pain transmission in mice. Here we investigated Tac4 mRNA expression in stress and pain-related regions and its involvement in chronic restraint stress-evoked behavioral changes and pain using Tac4 gene-deleted ( Tac4 -/- ) mice compared to C57Bl/6 wildtypes (WT). Tac4 mRNA was detected by in situ hybridization RNAscope technique. Touch sensitivity was assessed by esthesiometry, cold tolerance by paw withdrawal latency from 0°C water. Anxiety was evaluated in the light–dark box (LDB) and open field test (OFT), depression-like behavior in the tail suspension test (TST). Adrenal and thymus weights were measured at the end of the experiment. We found abundant Tac4 expression in the hypothalamic–pituitary–adrenal axis, but Tac4 mRNA was also detected in the hippocampus, amygdala, somatosensory and piriform cortices in mice, and in the frontal regions and the amygdala in humans. In Tac4 -/- mice of both sexes, stress-induced mechanical, but not cold hyperalgesia was significantly decreased compared to WTs. Stress-induced behavioral alterations were mild or absent in male WT animals, while significant changes of these parameters could be detected in females. Thymus weight decrease can be observed in both sexes. Higher baseline anxiety and depression-like behaviors were detected in male but not in female HK-1-deficient mice, highlighting the importance of investigating both sexes in preclinical studies. We provided the first evidence for the potent nociceptive and stress regulating effects of HK-1 in chronic restraint stress paradigm. Identification of its targets might open new perspectives for therapy of stress-induced pain.

With the advancement of molecular oncology, numerous new opportunities are available for the effective and efficient treatment of patients diagnosed with childhood brain tumors. This includes gene panel analysis aiding personalized treatment used in clinical trials, and the application of targeted therapy independent of tissue type (tumor agnostic therapy). Most personalized therapies inhibit certain kinases. In our review, we present the modern pathological diagnosis of childhood brain tumors, as well as the complex intracellular regulation of signal transduction pathways important from the point of view of clinical practice, and we describe their further targets defined on the basis of pharmacological characteristics of the pathway, based on international and our own results. Despite common mutations affecting kinases, personalized therapy is not available in many types of tumors. Through the example of childhood brain tumors, we demonstrate the expected future therapeutic significance of tyrosine kinases.

Glioblastomas are the most common IDH-wildtype adult high-grade gliomas, frequently harboring mutations in the TERT gene promoter ( pTERT ) and utilizing the subsequent telomerase overexpression for telomere length maintenance. However, some rare cases show loss of ATRX and use alternative mechanisms of telomere lengthening. In this study, we performed the first complex genomic analysis specifically concentrating on the latter subgroup. Comprehensive genomic profiling of 12 ATRX-deficient and 13 ATRX-intact IDH-wildtype adult high-grade gliomas revealed that ATRX and pTERT mutations are mutually exclusive. DNMT3A alterations were confined to ATRX-deficient, while PTEN mutations to ATRX-intact cases. RAS–MAPK pathway alterations, including NF1 mutations, were more characteristic in the ATRX-deficient group. Variants of genes related to homologous recombination repair showed different patterns of affected genes. Two ATRX-deficient tumors with high tumor mutational burden and mismatch repair deficiency were found. One of these contained a novel fusion involving the NTRK2 and LRRFIP2 genes, while the other showed loss of MSH2 and MSH6 without genetic alterations in the encoding genes suggesting an epigenetic background. Genetic characteristics of ATRX-deficient IDH-wildtype adult high-grade gliomas suggest that these tumors are particularly intriguing targets of potential future therapeutic interventions including immunotherapies combined with MAPK pathway inhibition and DNA repair inhibitors.

Oral squamous cell carcinoma (OSCC) is a common cancer with poor prognosis. Transient Receptor Potential Ankyrin 1 (TRPA1) and Vanilloid 1 (TRPV1) receptors are non-selective cation channels expressed on primary sensory neurons and epithelial and immune cells. TRPV1 mRNA and immunopositivity, as well as TRPA1-like immunoreactivity upregulation, were demonstrated in OSCC, but selectivity problems with the antibodies still raise questions and their functional relevance is unclear. Therefore, here, we investigated TRPA1 and TRPV1 expressions in OSCC and analyzed their functions. TRPA1 and TRPV1 mRNA were determined by RNAscope in situ hybridization and qPCR. Radioactive 45Ca2+ uptake and ATP-based luminescence indicating cell viability were measured in PE/CA-PJ41 cells in response to the TRPA1 agonist allyl-isothiocyanate (AITC) and TRPV1 agonist capsaicin to determine receptor function. Both TRPA1 and TRPV1 mRNA are expressed in the squamous epithelium of the human oral mucosa and in PE/CA-PJ41 cells, and their expressions are significantly upregulated in OSCC compared to healthy mucosa. TRPA1 and TRPV1 activation (100 µM AITC, 100 nM capsaicin) induced 45Ca2+-influx into PE/CA-PJ41 cells. Both AITC (10 nM–5 µM) and capsaicin (100 nM–45 µM) reduced cell viability, reaching significant decrease at 100 nM AITC and 45 µM capsaicin. We provide the first evidence for the presence of non-neuronal TRPA1 receptor in the OSCC and confirm the expression of TRPV1 channel. These channels are functionally active and might regulate cancer cell viability.

There is growing body of evidence supporting the role of germline mutations in the pathogenesis of pediatric central nervous system (CNS) tumors, and the widespread use of next-generation sequencing (NGS) panels facilitates their detection. Variants of the MUTYH gene are increasingly recognized as suspected germline background of various extraintestinal malignancies, besides their well-characterized role in the polyposis syndrome associated with biallelic mutations. Using a multigene NGS panel (Illumina TruSight Oncology 500), we detected one H3 G34V- and one H3 K27M-mutant pediatric high-grade diffuse glioma, in association with c.1178G>A (p.G393D) and c.916C>T (p.R306C) MUTYH variants, respectively. Both MUTYH mutations were germline, heterozygous and inherited, according to the subsequent genetic testing of the patients and their first-degree relatives. In the H3 K27M-mutant glioma, amplifications affecting the 4q12 region were also detected, in association with KDR-PDGFRA, KIT-PDGFRA, and KDR-CHIC2 fusions, previously unreported in this entity. Among 47 other CNS tumors of various histological types tested with the same NGS panel in our institution, only one adult glioblastoma harbored MUTYH mutation. Together with a single previous report, our data raises the possibility of an association between germline MUTYH mutations and CNS malignancies, particularly in pediatric histone H3-mutant gliomas.

Primary central nervous system (CNS) tumors are associated with dismal outcome and very poor survival rates. Comprehensive genomic profiling (CGP) approaches may contribute to the deeper understanding of the pathogenesis of these entities and can be harnessed to reveal potential therapeutic targets and offer personalized treatment options in patients with CNS tumors. Diagnostic samples of 42 patients with primary CNS tumors were investigated using Illumina TruSight Oncology 500 assay. Single nucleotide variants (SNV), small insertions and deletions, copy number alterations and gene fusions were analyzed along with microsatellite instability status and the tumor mutation burden. Actionable variants were determined using the Qiagen Clinical Insight and PierianDx software. Potentially targetable genomic alterations were identified in 69.0% (29/42) of the patients. Pathogenic SNVs, insertions and deletions were detected in 66.7% (28/42) of the patients with TP53, NTRK3 and FAT1 genes representing the most frequently affected loci. Twenty gene amplifications (copy number > 5) were uncovered in 19.0% patients, with MYC, PDGFRA and KIT genes being the most frequently revealed alterations. Gene fusions were detected in 10 (23.8%) patients. BRAF fusions, ETV5-DGKG translocations (4.4%), and NTRK3 fusions were detected in 9.5%, 4.4% and 2.4% of the cases, respectively. NGS revealed a novel BRAF::PID1gene fusion in a sample diagnosed with pilocytic astrocytoma. All but one patient was characterized by low tumor mutational burden and none of the analyzed patients displayed microsatellite-instability. Eleven patients (26.2%) have received targeted therapy based on their molecular profile. Trametinib, dasatinib and cabozantinib were the most commonly used agents. Partial remission or stable disease has been observed in patients receiving targeted therapies for at least 6 months. In our study, routine application of the Illumina TruSight Oncology 500 CPG approach led to the identification of targetable alterations in 69.0% of the patients with 26.2% of the patients receiving targeted therapy.

Medication-Related Osteonecrosis of the Jaws (MRONJ) is a difficult-to-treat complication of the therapy of osteoporosis and some malignancies cured with bisphosphonates and antiresorptive drugs. The pathomechanism is unclear, but there is increasing observation that Actinomyces infection may play a role in its development and progression. The aim of our study was to demonstrate that histological examination using a validated triple staining procedure for Actinomyces bacteria strains can detect a high rate of Actinomyces infection in patient's samples with MRONJ. 112 previously hematoxylin-eosin (HE) stained samples submitted with the clinical diagnosis of MRONJ were re-evaluated histologically using an appropriate triple special staining validated for the identification of Actinomyces infection. During the first evaluation, when pathologists did not specifically look for Actinomyces, only 8.93% of the samples were reported as positive. In contrast, re-evaluation with triple staining provided a yield of 93.7% positive samples, therefore, we suggest the triple special staining to be standard in MRONJ histology evaluation. These results show that if the clinician suspects Actinomyces infection and brings this to the attention of the pathologist, it could significantly increase the number of correct diagnoses. It serves as an aid for clinicians in therapeutic success of MRONJ by selecting a long-term adequate antibiotic medication which is suitable for the elimination of actinomyces infection.

Background: Nonsteroidal anti-inflammatory drugs (NSAIDs) induce significant damage to the small intestine, which is accompanied by changes in intestinal bacteria (dysbiosis) and bile acids. However, it is still a question of debate whether besides mucosal inflammation also other factors, such as direct antibacterial effects or delayed peristalsis, contribute to NSAID-induced dysbiosis. Here we aimed to assess whether ketorolac, an NSAID lacking direct effects on gut bacteria, has any significant impact on intestinal microbiota and bile acids in the absence of mucosal inflammation. We also addressed the possibility that ketorolac-induced bacterial and bile acid alterations are due to a delay in gastrointestinal (GI) transit. Methods: Vehicle or ketorolac (1, 3 and 10 mg/kg) were given to rats by oral gavage once daily for four weeks, and the severity of mucosal inflammation was evaluated macroscopically, histologically, and by measuring the levels of inflammatory proteins and claudin-1 in the distal jejunal tissue. The luminal amount of bile acids was measured by liquid chromatography-tandem mass spectrometry, whereas the composition of microbiota by sequencing of bacterial 16S rRNA. GI transit was assessed by the charcoal meal method. Results: Ketorolac up to 3 mg/kg did not cause any signs of mucosal damage to the small intestine. However, 3 mg/kg of ketorolac induced dysbiosis, which was characterized by a loss of families belonging to Firmicutes ( Paenibacillaceae , Clostridiales Family XIII , Christensenellaceae ) and bloom of Enterobacteriaceae . Ketorolac also changed the composition of small intestinal bile by decreasing the concentration of conjugated bile acids and by increasing the amount of hyodeoxycholic acid (HDCA). The level of conjugated bile acids correlated negatively with the abundance of Erysipelotrichaceae , Ruminococcaceae , Clostridiaceae 1 , Muribaculaceae , Bacteroidaceae , Burkholderiaceae and Bifidobacteriaceae . Ketorolac, under the present experimental conditions, did not change the GI transit. Conclusion: This is the first demonstration that low-dose ketorolac disturbed the delicate balance between small intestinal bacteria and bile acids, despite having no significant effect on intestinal mucosal integrity and peristalsis. Other, yet unidentified, factors may contribute to ketorolac-induced dysbiosis and bile dysmetabolism.

There is some recent evidence that cardiac ischemia/reperfusion (I/R) injury induces intestinal damage within days, which contributes to adverse cardiovascular outcomes after myocardial infarction. However, it is not clear whether remote gut injury has any detectable early signs, and whether different interventions aiming to reduce cardiac damage are also effective at protecting the intestine. Previously, we found that chronic treatment with rofecoxib, a selective inhibitor of cyclooxygenase-2 (COX-2), limited myocardial infarct size to a comparable extent as cardiac ischemic preconditioning (IPC) in rats subjected to 30-min coronary artery occlusion and 120-min reperfusion. In the present study, we aimed to analyse the early intestinal alterations caused by cardiac I/R injury, with or without the above-mentioned infart size-limiting interventions. We found that cardiac I/R injury induced histological changes in the small intestine within 2 h, which were accompanied by elevated tissue level of COX-2 and showed positive correlation with the activity of matrix metalloproteinase-2 (MMP-2), but not of MMP-9 in the plasma. All these changes were prevented by rofecoxib treatment. By contrast, cardiac IPC failed to reduce intestinal injury and plasma MMP-2 activity, although it prevented the transient reduction in jejunal blood flow in response to cardiac I/R. Our results demonstrate for the first time that rapid development of intestinal damage follows cardiac I/R, and that two similarly effective infarct size-limiting interventions, rofecoxib treatment and cardiac IPC, have different impacts on cardiac I/R-induced gut injury. Furthermore, intestinal damage correlates with plasma MMP-2 activity, which may be a biomarker for its early diagnosis.

The Tac4 gene-derived hemokinin-1 (HK-1) is present in pain-related regions and activates the tachykinin NK1 receptor, but with binding site and signaling pathways different from Substance P (SP). NK1 receptor is involved in nociception, but our earlier data showed that it has no role in chronic neuropathic hyperalgesia, similarly to SP. Furthermore, NK1 antagonists failed in clinical trials as analgesics due to still unknown reasons. Therefore, we investigated the role of HK-1 in pain conditions of distinct mechanisms using genetically modified mice. Chronic neuropathic mechanical and cold hyperalgesia after partial sciatic nerve ligation (PSL) were determined by dynamic plantar aesthesiometry and withdrawal latency from icy water, motor coordination on the accelerating Rotarod. Peripheral nerve growth factor (NGF) production was measured by ELISA, neuronal and glia cell activation by immunohistochemistry in pain-related regions. Acute somatic and visceral chemonocifensive behaviors were assessed after intraplantar formalin or intraperitoneal acetic-acid injection, respectively. Resiniferatoxin-induced inflammatory mechanical and thermal hyperalgesia by aesthesiometry and increasing temperature hot plate. Chronic neuropathic mechanical and cold hypersensitivity were significantly decreased in HK-1 deficient mice. NGF level in the paw homogenates of intact mice were significantly lower in case of HK-1 deletion. However, it significantly increased under neuropathic condition in contrast to wildtype mice, where the higher basal concentration did not show any changes. Microglia, but not astrocyte activation was observed 14 days after PSL in the ipsilateral spinal dorsal horn of wildtype, but not HK-1-deficient mice. However, under neuropathic conditions, the number of GFAP-positive astrocytes was significantly smaller in case of HK-1 deletion. Acute visceral, but not somatic nocifensive behavior, as well as neurogenic inflammatory mechanical and thermal hypersensitivity were significantly reduced by HK-1 deficiency similarly to NK1, but not to SP deletion. We provide evidence for pro-nociceptive role of HK-1, via NK1 receptor activation in acute inflammation models, but differently from SP-mediated actions. Identification of its targets and signaling can open new directions in pain research.