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Canines can identify prostate cancer with high accuracy by smelling volatile organic compounds (VOCs) in urine. Previous studies have identified VOC biomarkers for prostate cancer utilizing solid phase microextraction (SPME) gas chromatography–mass spectrometry (GC-MS) but have not assessed the ability of VOCs to distinguish aggressive cancers. Additionally, previous investigations have utilized murine models to identify biomarkers but have not determined if the results are translatable to humans. To address these challenges, urine was collected from mice with prostate cancer and men undergoing prostate cancer biopsy and VOCs were analyzed by SPME GC-MS. Prior to analysis, SPME fibers/arrows were compared, and the fibers had enhanced sensitivity toward VOCs with a low molecular weight. The analysis of mouse urine demonstrated that VOCs could distinguish tumor-bearing mice with 100% accuracy. Linear discriminant analysis of six VOCs in human urine distinguished prostate cancer with sensitivity = 75% and specificity = 69%. Another panel of seven VOCs could classify aggressive cancer with sensitivity = 78% and specificity = 85%. These results show that VOCs have moderate accuracy in detecting prostate cancer and a superior ability to stratify aggressive tumors. Furthermore, the overlap in the structure of VOCs identified in humans and mice shows the merit of murine models for identifying biomarker candidates.

Background and Hypothesis:Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with a very poor survival rate. The PI3K pathway has been studied extensively in cancer cells which has led to the creation of PI3K inhibitor drugs. To explore an uncharted and counterintuitive use of the PI3K pathway, PI3K was activated in mesenchymal stem cells (MSCs) and the action of their conditioned medium (CM) on PDAC was examined. We hypothesized that PI3K-activated MSC-derived CM is capable of suppressing tumor growth and migration. Project Methods:To generate the treatment conditioned media, we first isolated and grew MSCs in DMEM + 10% serum and added 50 μM of PI3K activator (YS49) and incubated for 24 hours. Afterwards, we exchanged the media with fresh DMEM + 10% serum and incubated for another 24 hours. Next, we hyper-centrifuged the media to extract the conditioned media and used the media to treat PANC-1, PA03C, ASPC-1, PANC10.05, and PANC198 pancreatic cancer cell lines. In vitro viability and migration assays were conducted on these five treated cell lines. The regulatory mechanism was evaluated using Western blotting, immunoprecipitation, and ELISAs. Results:PI3K-activated MSC-derived CM reduced the proliferation (N=5) and migration (N=2) of PDAC (p < 0.01). MSC CM was enriched with polyubiquitin C (Ubc), which acted as an extracellular tumor-suppressing protein by interacting with CXCR4. While the anti-tumor efficacy of Ubc differed depending on the CXCR4 expression level of PDAC, MSC CM was commonly effective in the tested cell lines and tissues (N=5). Conclusion and potential impact:Collectively, this study demonstrated that MSC CM can be converted into a tumor-suppressing agent by the activation of PI3K, and a Ubc-CXCR4 regulatory axis is at least in part responsible. The use of MSC CM treatment can potentially serve as an alternative or supplemental treatment to current chemotherapeutic agents in treating pancreatic cancer without inducing life-threatening side effects.

To understand the direct impact of bradykinin in autonomic control of circulation through baroreflex afferent pathway.The mean arterial pressure (MAP) was monitored while bradykinin and its agonists were applied via nodose (NG) microinjection, the expression of bradykinin receptors (BRs) in the NG (1Under physiological condition, bradykinin-induced dose- and estrogen-dependent reductions of MAP with lower estimated ECThese datasets have demonstrated for the first time that bradykinin mediates direct activation of baroreflex afferent function to trigger estrogen-dependent depressor response, which is due mainly to the direct activation/neuroexcitation of female-specific myelinated Ah-type baroreceptor neurons leading to a sexual dimorphism in parasympathetic domination of blood pressure regulation via activation of B

Tumor cells regulate their progression not only by the factors within their cell bodies but also by the secretome they produce and secrete. While their secretome significantly alters the fate of tumor cells themselves, they also regulate the growth of surrounding cells including both companion cancer and non-cancer cells. Tumor cell secretome consists of varying molecules that have been reported mostly tumor-promotive. Furthermore, their pro-tumor capability is enhanced by the application of chemotherapeutic agents. However, multiple lines of emerging evidence suggest that the tumor cell secretome can be tumor-suppressive in response to paracrine and endocrine signaling. This review introduces both tumor-promotive and tumor-suppressive secretomes, focusing on multi-tasking proteins in the intracellular and extracellular domains. We describe tumorigenic signaling that governs the nature of the tumor cell secretome and discuss the possibility of inducing tumor-suppressive proteomes as a novel option for cancer treatment. We evaluated the counterintuitive procedure to generate tumor-suppressive proteomes from a unique type of tumor-modifying cells, which are named “induced tumor-suppressing cells” (iTSCs).

Background: Calsequestrins (Casqs), comprising the Casq1 and Casq2 isoforms, buffer Ca 2+ and regulate its release in the sarcoplasmic reticulum of skeletal and cardiac muscle, respectively. Human inherited diseases associated with mutations in CASQ1 or CASQ2 include malignant hyperthermia/environmental heat stroke (MH/EHS) and catecholaminergic polymorphic ventricular tachycardia. However, patients with an MH/EHS event often experience arrhythmia for which the underlying mechanism remains unknown. Methods: Working hearts from conventional ( Casq1 -KO) and cardiac-specific ( Casq1 -CKO) Casq1 knockout mice were monitored in vivo and ex vivo by ECG and electric mapping, respectively. MH was induced by 2% isoflurane and treated intraperitoneally with dantrolene. Time-lapse imaging was used to monitor intracellular Ca 2+ activity in isolated mouse cardiomyocytes or neonatal rat ventricular myocytes with knockdown, overexpression, or truncation of the Casq1 gene. Conformational change in both Casqs was determined by cross-linking Western blot analysis. Results: Like patients with MH/EHS, Casq1 -KO and Casq1 -CKO mice had faster basal heart rate and ventricular tachycardia on exposure to 2% isoflurane, which could be relieved by dantrolene. Basal sinus tachycardia and ventricular ectopic electric triggering also occurred in Casq1 -KO hearts ex vivo. Accordingly, the ventricular cardiomyocytes from Casq1 -CKO mice displayed dantrolene-sensitive increased Ca 2+ waves and diastole premature Ca 2+ transients/oscillations on isoflurane. Neonatal rat ventricular myocytes with Casq1-knockdown had enhanced spontaneous Ca 2+ sparks/transients on isoflurane, whereas cells overexpressing Casq1 exhibited decreased Ca 2+ sparks/transients that were absent in cells with truncation of 9 amino acids at the C terminus of Casq1. Structural evaluation showed that most of the Casq1 protein was present as a polymer and physically interacted with ryanodine receptor-2 in the ventricular sarcoplasmic reticulum. The Casq1 isoform was also expressed in human myocardium. Mechanistically, exposure to 2% isoflurane or heating at 41 °C induced Casq1 oligomerization in mouse ventricular and skeletal muscle tissues, leading to a reduced Casq1/ryanodine receptor-2 interaction and increased ryanodine receptor-2 activity in the ventricle. Conclusions: Casq1 is expressed in the heart, where it regulates sarcoplasmic reticulum Ca 2+ release and heart rate. Casq1 deficiency independently causes MH/EHS-like ventricular arrhythmia by trigger-induced Casq1 oligomerization and a relief of its inhibitory effect on ryanodine receptor-2–mediated Ca 2+ release, thus revealing a new inherited arrhythmia and a novel mechanism for MH/EHS arrhythmogenesis.

It is increasingly recognized that a single protein can have multiple, sometimes paradoxical, roles in cell functions as well as pathological conditions depending on its cellular locations. Here we report that moesins (MSNs) in the intracellular and extracellular domains present opposing roles in pro-tumorigenic signaling in breast cancer cells. Using live cell imaging with fluorescence resonance energy transfer (FRET)- and green fluorescent protein (GFP)-based biosensors, we investigated the molecular mechanism underlying the cellular location-dependent effect of MSN on Src and β-catenin signaling in MDA-MB-231 breast cancer cells. Inhibition of intracellular MSN decreased the activities of Src and FAK, whereas overexpression of intracellular MSN increased them. By contrast, extracellular MSN decreased the activities of Src, FAK, and RhoA, as well as β-catenin translocation to the nucleus. Consistently, Western blotting and MTT-based analysis showed that overexpression of intracellular MSN elevated the expression of oncogenic genes, such as p-Src, β-catenin, Lrp5, MMP9, Runx2, and Snail, as well as cell viability, whereas extracellular MSN suppressed them. Conditioned medium derived from MSN-overexpressing mesenchymal stem cells or osteocytes showed the anti-tumor effects by inhibiting the Src activity and β-catenin translocation to the nucleus as well as the activities of FAK and RhoA and MTT-based cell viability. Conditioned medium derived from MSN-inhibited cells increased the Src activity, but it did not affect the activities of FAK and RhoA. Silencing CD44 and/or FN1 in MDA-MB-231 cells blocked the suppression of Src activity and β-catenin accumulation in the nucleus by extracellular MSN. Collectively, the results suggest that cellular location-specific MSN is a strong regulator of Src and β-catenin signaling in breast cancer cells, and that extracellular MSN exerts tumor-suppressive effects via its interaction with CD44 and FN1.

Female-specific subpopulation of myelinated Ah-type baroreceptor neurons (BRNs) in nodose ganglia is the neuroanatomical base of sexual-dimorphic autonomic control of blood pressure regulation, and KCa1.1 is a key player in modulating the neuroexcitation in nodose ganglia. In this study we investigated the exact mechanisms underlying KCa1.1-mediated neuroexcitation of myelinated Ah-type BRNs in the presence or absence of estrogen. BRNs were isolated from adult ovary intact (OVI) or ovariectomized (OVX) female rats, and identified electrophysiologically and fluorescently. Action potential (AP) and potassium currents were recorded using whole-cell recording. Consistently, myelinated Ah-type BRNs displayed a characteristic discharge pattern and significantly reduced excitability after OVX with narrowed AP duration and faster repolarization largely due to an upregulated iberiotoxin (IbTX)-sensitive component; the changes in AP waveform and repetitive discharge of Ah-types from OVX female rats were reversed by G1 (a selective agonist for estrogen membrane receptor GPR30, 100 nM) and/or IbTX (100 nM). In addition, the effect of G1 on repetitive discharge could be completely blocked by G15 (a selective antagonist for estrogen membrane receptor GPR30, 3 μM). These data suggest that estrogen deficiency by removing ovaries upregulates KCa1.1 channel protein in Ah-type BRNs, and subsequently increases AP repolarization and blunts neuroexcitation through estrogen membrane receptor signaling. Intriguingly, this upregulated KCa1.1 predicted electrophysiologically was confirmed by increased mean fluorescent intensity that was abolished by estrogen treatment. These electrophysiological findings combined with immunostaining and pharmacological manipulations reveal the crucial role of KCa1.1 in modulation of neuroexcitation especially in female-specific subpopulation of myelinated Ah-type BRNs and extend our current understanding of sexual dimorphism of neurocontrol of BP regulation.

Purpose: To develop a novel treatment option for Chondrosarcoma (CS) and inflammatory arthritis, we evaluated a counterintuitive approach of activating tumorigenic and inflammatory signaling for generating joint-protective proteomes. Methods: We employed mesenchymal stem cells and chondrocytes to generate chondroprotective proteomes by activating PI3K signaling and the administration of TNFα. The efficacy of the proteomes was examined using human and mouse cell lines as well as a mouse model of CS. The regulatory mechanism was analyzed using mass spectrometry-based whole-genome proteomics. Results: While tumor progression and inflammatory responses were promoted by activating PI3K signaling and the administration of TNFα to CS cells and chondrocytes, those cells paradoxically generated a chondroprotective conditioned medium (CM). The application of CM downregulated tumorigenic genes in CS cells and TNFα and MMP13 in chondrocytes. Mechanistically, Hsp90ab1 was enriched in the chondroprotective CM, and it immunoprecipitated GAPDH. Extracellular GAPDH interacted with L1CAM and inhibited tumorigenic behaviors, whereas intracellular GAPDH downregulated p38 and exerted anti-inflammatory effects. Conclusions: We demonstrated that the unconventional approach of activating oncogenic and inflammatory signaling can generate chondroprotective proteomes. The role of Hsp90ab1 and GAPDH differed in their locations and they acted as the uncommon protectors of the joint tissue from tumor and inflammatory responses.

Mechanical loading to the bone is known to be beneficial for bone homeostasis and for suppressing tumor-induced osteolysis in the loaded bone. However, whether loading to a weight-bearing hind limb can inhibit distant tumor growth in the brain is unknown. We examined the possibility of bone-to-brain mechanotransduction using a mouse model of a brain tumor by focusing on the response to Lrp5-mediated Wnt signaling and dopamine in tumor cells. The results revealed that loading the tibia with elevated levels of tyrosine hydroxylase, a rate-limiting enzyme in dopamine synthesis, markedly reduced the progression of the brain tumors. The simultaneous application of fluphenazine (FP), an antipsychotic dopamine modulator, enhanced tumor suppression. Dopamine and FP exerted antitumor effects through the dopamine receptors DRD1 and DRD2, respectively. Notably, dopamine downregulated Lrp5 via DRD1 in tumor cells. A cytokine array analysis revealed that the reduction in CCN4 was critical for loading-driven, dopamine-mediated tumor suppression. The silencing of Lrp5 reduced CCN4, and the administration of CCN4 elevated oncogenic genes such as MMP9, Runx2, and Snail. In summary, this study demonstrates that mechanical loading regulates dopaminergic signaling and remotely suppresses brain tumors by inhibiting the Lrp5-CCN4 axis via DRD1, indicating the possibility of developing an adjuvant bone-mediated loading therapy.

Aim To understand why autonomic failures, a common non‐motor symptom of Parkinson's disease (PD), occur earlier than typical motor disorders. Methods Vagal application of DOPAL (3,4‐dihydroxyphenylacetaldehyde) to simulate PD‐like autonomic dysfunction and understand the connection between PD and cardiovascular dysfunction. Molecular and morphological approaches were employed to test the time‐dependent alternation of α‐synuclein aggregation and the ultrastructure changes in the heart and nodose (NG)/nucleus tractus solitarius (NTS). Results Blood pressure (BP) and baroreflex sensitivity of DOPAL‐treated rats were significantly reduced accompanied with a time‐dependent change in orthostatic BP, consistent with altered echocardiography and cardiomyocyte mitochondrial ultrastructure. Notably, time‐dependent and collaborated changes in Mon‐/Tri‐α‐synuclein were paralleled with morphological alternation in the NG and NTS. Conclusion These all demonstrate that early autonomic dysfunction mediated by vagal application of DOPAL highly suggests the plausible etiology of PD initiated from peripheral, rather than central site. It will provide a scientific basis for the prevention and early diagnosis of PD., Application of DOPAL on Vagus causes significant accumulation of α‐Syn monomers to form toxic oligomers that could be transported to the heart TA and NG‐NTS by axon flow

Rationale: The progression of cancer cells depends on the soil and building an inhibitory soil might be a therapeutic option. We previously created tumor-suppressive secretomes by activating Wnt signaling in MSCs. Here, we examined whether the anti-tumor secretomes can be produced from tumor cells. Methods: Wnt signaling was activated in tumor cells by overexpressing β-catenin or administering BML284, a Wnt activator. Their conditioned medium (CM) was applied to cancer cells or tissues, and the effects of CM were evaluated. Tumor growth in the mammary fat pad and tibia in C57BL/6 female mice was also evaluated through μCT imaging and histology. Whole-genome proteomics analysis was conducted to determine and characterize novel tumor-suppressing proteins, which were enriched in CM. Results: The overexpression of β-catenin or the administration of BML284 generated tumor-suppressive secretomes from breast, prostate and pancreatic cancer cells. In the mouse model, β-catenin-overexpressing CM reduced tumor growth and tumor-driven bone destruction. This inhibition was also observed with BML284-treated CM. Besides p53 and Trail, proteomics analysis revealed that CM was enriched with enolase 1 (Eno1) and ubiquitin C (Ubc) that presented notable tumor-suppressing actions. Importantly, Eno1 immunoprecipitated CD44, a cell-surface adhesion receptor, and its silencing suppressed Eno1-driven tumor inhibition. A pan-cancer survival analysis revealed that the downregulation of MMP9, Runx2 and Snail by CM had a significant impact on survival outcomes (p < 0.00001). CM presented a selective inhibition of tumor cells compared to non-tumor cells, and it downregulated PD-L1, an immune escape modulator. Conclusions: The tumor-suppressive secretome can be generated from tumor cells, in which β-catenin presented two opposing roles, as an intracellular tumor promoter in tumor cells and a generator of extracellular tumor suppressor in CM. Eno1 was enriched in CM and its interaction with CD44 was involved in Eno1's anti-tumor action. Besides presenting a potential option for treating primary cancers and metastases, the result indicates that aggressive tumors may inhibit the growth of less aggressive tumors via tumor-suppressive secretomes.

Wnt signaling plays a critical role in loading-driven bone formation and bone homeostasis, whereas its activation in cancer cells promotes their progression. Currently, major research efforts in cancer treatment have been directed to the development of Wnt inhibitors. Recent studies on tumor-bone interactions, however, presented multiple lines of evidence that support a tumor-suppressive role of Lrp5, a Wnt co-receptor, and β-catenin, in Wnt signaling. This review describes the action of Wnt signaling as a double-edged sword in the bone microenvironment and suggests the possibility of a novel option for protecting bone from cancer.

Hydrogen sulfide (H2S), which is closely related to various cardiovascular disorders, lowers blood pressure (BP), but whether this action is mediated via the modification of baroreflex afferent function has not been elucidated. Therefore, the current study aimed to investigate the role of the baroreflex afferent pathway in H2S-mediated autonomic control of BP regulation. The results showed that baroreflex sensitivity (BRS) was increased by acute intravenous NaHS (a H2S donor) administration to renovascular hypertensive (RVH) and control rats. Molecular expression data also showed that the expression levels of critical enzymes related to H2S were aberrantly downregulated in the nodose ganglion (NG) and nucleus tractus solitarius (NTS) in RVH rats. A clear reduction in BP by the microinjection of NaHS or L-cysteine into the NG was confirmed in both RVH and control rats, and a less dramatic effect was observed in model rats. Furthermore, the beneficial effects of NaHS administered by chronic intraperitoneal infusion on dysregulated systolic blood pressure (SBP), cardiac parameters, and BRS were verified in RVH rats. Moreover, the increase in BRS was attributed to activation and upregulation of the ATP-sensitive potassium (KATP) channels Kir6.2 and SUR1, which are functionally expressed in the NG and NTS. In summary, H2S plays a crucial role in the autonomic control of BP regulation by improving baroreflex afferent function due at least in part to increased KATP channel expression in the baroreflex afferent pathway under physiological and hypertensive conditions.

Large conductance of Ca2+-activated K+ channel (KCa1.1) plays an inhibitory role in neuroexcitation. However, the expression of KCNMB4/β4-subunit in the nodose ganglia (NG) and nucleus tractus solitarius (NTS), and its effect and regulation on baroreflex afferent function at post-transcriptional level of female rats remains unknown. Here, we demonstrated that the expression of β4-subunit encoded by KCNMB4 was significantly lower in females vs. males and ovariectomized (OVX) rats in the NG. Although all baroreceptor neurons (BRNs) expressed β4-subunit, altered discharge characteristics were only observed in Ah-type neurons after ovariectomy. Notably, the decreased excitability of Ah-types was restored by paxilline and further enhanced by iberiotoxin. The consistent changes were observed in excitatory post-synaptic currents. The level of miR-504 was higher in females, which was predicted to bind to the 3'UTR of KCNMB4. In consistent, an inverse expression pattern between miR-504 and KCNMB4 was observed in baroreflex afferents. The paxilline-sensitive β4-subunits is less in Ah-types and up-regulated by ovariectomy. These data indicated that KCa1.1 β4-subunit is the key regulator in neuroexcitation of Ah-types and sexual-dimorphism in baroreflex afferent function through estrogen-dependent inhibition of KCNMB4 expression via miR-504.

Silent myocardial infarction (MI) is critical for clinical practice with increasing risk for women and the cause remains a medical mystery. Upon the discovery of female-specific Ah-type baroreceptor neurons (BRNs), we hypothesize that glutamate mediates depressor response through afferent-specific expression of particular glutamate receptors (mGluRs) leading descending inhibition of cardiac nociception. In vivo, tail-flick reflex and electromyography were assessed to evaluate glutamate-mediated blood pressure regulation, peripheral and cardiac nociception. The results showed that glutamate decreased mean arterial pressure (MAP) and increased peripheral nociception. Interestingly, glutamate-mediated capsaicin-induced cardiac nociception was strongly reduced in female rats compared with males. Furthermore, Nodose (NG) microinjection of mGluR7 agonist significantly increased MAP in males and slightly decreased that in females. Even though mGluR8 direct activation intensified baroreceptor activation, the sensitivity was similar between sexes. In vitro, the expression profiles of mGluRs were investigated using Western blot and identified BRNs using single-cell qRT-PCR under ischemic conditions. Glutamate in serum, NG and nucleus tractus solitary (NTS) was raised significantly in the model rats of both sexes vs. sham-controls. Female-specific expression of mGluR7 in the baroreflex afferent pathway, especially higher expression in Ah-type BRNs, contributes significantly to cardiac analgesia, which may explain that the pathogenesis of silent MI occurs mainly in female patients. Therefore, higher expression of mGluR7 in female-specific subpopulation of Ah-type BRNs plays a critical role in cardiac analgesia and peripheral nociception.

Recent studies suggest that melatonin (Mel) plays an important role in the regulation of blood pressure (BP) via the aortic baroreflex pathway. In this study, we investigated the interaction between the baroreflex afferent pathway and Mel-mediated BP regulation in rats under physiological and hypertensive conditions. Mel (0.1, 0.3, and 1.0 mg/mL) was microinjected into the nodose ganglia (NG) of rats. We showed that Mel-induced reduction of mean arterial pressure in female rats was significantly greater than that in male and in ovariectomized rats under physiological condition. Consistently, the expression of Mel receptors (MTNRs) in the NG of female rats was significantly higher than that of males. In L-NAME-induced hypertensive and spontaneously hypertensive rat models, MTNRs were upregulated in males but downregulated in female models. Interestingly, Mel-induced BP reduction was found in male hypertensive models. In whole-cell recording from identified baroreceptor neurons (BRNs) in female rats, we found that Mel (0.1 μM) significantly increased the excitability of a female-specific subpopulation of Ah-type BRNs by increasing the Na

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Background: Bone is a frequent site of metastases from breast cancer, but existing therapeutic options are not satisfactory. Although osteoblasts have active roles in cancer progression by assisting the vicious bone-destructive cycle, we employed a counterintuitive approach of activating pro-tumorigenic Wnt signaling and examined the paradoxical possibility of developing osteoblast-derived tumor-suppressive, bone-protective secretomes. Methods: Wnt signaling was activated by the overexpression of Lrp5 and β-catenin in osteoblasts as well as a pharmacological agent (BML284), and the therapeutic effects of their conditioned medium (CM) were evaluated using in vitro cell cultures, ex vivo breast cancer tissues, and a mouse model of osteolysis. To explore the unconventional regulatory mechanism of the action of Wnt-activated osteoblasts, whole-genome proteomics analysis was conducted, followed by immunoprecipitation and gain- and loss-of-function assays. Results: While osteoblasts did not present any innate tumor-suppressing ability, we observed that the overexpression of Lrp5 and β-catenin in Wnt signaling made their CM tumor-suppressive and bone-protective. The growth of breast cancer cells and tissues was inhibited by Lrp5-overexpressing CM (Lrp5 CM), which suppressed mammary tumors and tumor-driven bone destruction in a mouse model. Lrp5 CM also inhibited the differentiation and maturation of bone-resorbing osteoclasts by downregulating NFATc1 and cathepsin K. The overexpression of Lrp5 upregulated osteopontin that enriched Hsp90ab1 (Hsp90 beta) and moesin (MSN) in Lrp5 CM. Hsp90ab1 and MSN are atypical tumor-suppressing proteins since they are multi-tasking, moonlighting proteins that promote tumorigenesis in tumor cells. Importantly, Hsp90ab1 immuno-precipitated latent TGFβ and inactivated TGFβ, whereas MSN interacted with CD44, a cancer stem-cell marker, as well as fibronectin 1, an ECM protein. Furthermore, Hsp90ab1 and MSN downregulated KDM3A that demethylated histones, together with PDL1 that inhibited immune responses. Conclusion: In contrast to inducing tumor-enhancing secretomes and chemoresistance in general by inhibiting varying oncogenic pathways in chemotherapy, this study presented the unexpected outcome of generation tumor-suppressive secretomes by activating the pro-tumorigenic Wnt pathway. The results shed light on the contrasting role of oncogenic signaling in tumor cells and osteoblast-derived secretomes, suggesting a counterintuitive option for the treatment of breast cancer-associated bone metastasis.

Osteocytes are the most abundant cells in bone, which is a frequent site of breast cancer metastasis. Here, we focused on Wnt signaling and evaluated tumor-osteocyte interactions. In animal experiments, mammary tumor cells were inoculated into the mammary fat pad and tibia. The role of Lrp5-mediated Wnt signaling was examined by overexpressing and silencing Lrp5 in osteocytes and establishing a conditional knockout mouse model. The results revealed that administration of osteocytes or their conditioned medium (CM) inhibited tumor progression and osteolysis. Osteocytes overexpressing Lrp5 or β-catenin displayed strikingly elevated tumor-suppressive activity, accompanied by downregulation of tumor-promoting chemokines and upregulation of apoptosis-inducing and tumor-suppressing proteins such as p53. The antitumor effect was also observed with osteocyte-derived CM that was pretreated with a Wnt-activating compound. Notably, silencing Lrp5 in tumors inhibited tumor progression, while silencing Lrp5 in osteocytes in conditional knockout mice promoted tumor progression. Osteocytes exhibited elevated Lrp5 expression in response to tumor cells, implying that osteocytes protect bone through canonical Wnt signaling. Thus, our results suggest that the Lrp5/β-catenin axis activates tumor-promoting signaling in tumor cells but tumor-suppressive signaling in osteocytes. We envision that osteocytes with Wnt activation potentially offer a novel cell-based therapy for breast cancer and osteolytic bone metastasis.