Noninvasive ultrasound stimulation to treat myocarditis through splenic neuro-immune regulation.

BACKGROUND: The cholinergic anti-inflammatory pathway (CAP) has been widely studied to modulate the immune response. Current stimulating strategies are invasive or imprecise. Noninvasive low-intensity pulsed ultrasound (LIPUS) has become increasingly appreciated for targeted neuronal modulation. However, its mechanisms and physiological role on myocarditis remain poorly defined. METHODS: The mouse model of experimental autoimmune myocarditis was established. Low-intensity pulsed ultrasound was targeted at the spleen to stimulate the spleen nerve. Under different ultrasound parameters, histological tests and molecular biology were performed to observe inflammatory lesions and changes in immune cell subsets in the spleen and heart. In addition, we evaluated the dependence of the spleen nerve and cholinergic anti-inflammatory pathway of low-intensity pulsed ultrasound in treating autoimmune myocarditis in mice through different control groups. RESULTS: The echocardiography and flow cytometry of splenic or heart infiltrating immune cells revealed that splenic ultrasound could alleviate the immune response, regulate the proportion and function of CD4+ Treg and macrophages by activating cholinergic anti-inflammatory pathway, and finally reduce heart inflammatory injury and improve cardiac remodeling, which is as effective as an acetylcholine receptor agonists GTS-21. Transcriptome sequencing showed significant differential expressed genes due to ultrasound modulation. CONCLUSIONS: It is worth noting that the ultrasound therapeutic efficacy depends greatly on acoustic pressure and exposure duration, and the effective targeting organ was the spleen but not the heart. This study provides novel insight into the therapeutic potentials of LIPUS, which are essential for its future application.

Apoptotic tumor cell-derived microparticles loading Napabucasin inhibit CSCs and synergistic immune therapy.

BACKGROUND: Cancer stem cells (CSCs) are crucial for the growth, metastasis, drug resistance, recurrence, and spread of tumors. Napabucasin (NAP) could effectively inhibit CSC, but its mechanism has not been fully explained. Additionally, NAP also has the drawbacks of poor water solubility and low utilization. Therefore, this study not only elaborated the new mechanism of NAP inhibiting CSCs, but also built NAP-loaded nanoprobes using apoptotic tumor-derived microparticles (TMPs) as carriers to combine diagnose and treat of colon cancer and lessen the adverse effects of NAP. RESULTS: The study discovered a new mechanism for NAP inhibiting tumors. NAP, in addition to inhibiting STAT3, may also inhibit STAT1, thereby inhibiting the expression of CD44, and the stemness of colon cancer. N3-TMPs@NAP was successfully synthesized, and it possessed a lipid bilayer with a particle size of 220.13 ± 4.52 nm, as well as strong tumor binding ability and anti-tumor effect in vitro. In static PET/CT imaging studies, the tumor was clearly visible and showed higher uptake after N3-TMPs@NAP injection than after oral administration. The average tumor volume and weight of the N3-TMPs@NAP group on day 14 of the treatment studies were computed to be 270.55 ± 107.59 mm3 and 0.30 ± 0.12 g, respectively. These values were significantly lower than those of the other groups. Additionally, N3-TMPs@NAP might prevent colon cancer from spreading to the liver. Furthermore, due to TMPs' stimulation of innate immunity, N3-TMPs@NAP might stimulate anti-tumor. CONCLUSIONS: As a combined diagnostic and therapeutic nanoprobe, N3-TMPs@NAP could successfully conduct PET/CT imaging, suppress CSCs, and synergistically stimulate anticancer immune responses. Additionally, this nanoprobe might someday be employed in clinical situations because TMPs for it can be produced from human tissue and NAP has FDA approval.

Ultrasound-targeted microbubble destruction promotes PDGF-primed bone mesenchymal stem cell transplantation for myocardial protection in acute Myocardial Infarction in rats.

BACKGROUND: Ultrasound-targeted microbubble destruction (UTMD) has emerged as a promising strategy for the targeted delivery of bone marrow mesenchymal stem cells (MSCs) to the ischemic myocardium. However, the limited migration capacity and poor survival of MSCs remains a major therapeutic barrier. The present study was performed to investigate the synergistic effect of UTMD with platelet-derived growth factor BB (PDGF-BB) on the homing of MSCs for acute myocardial infarction (AMI). METHODS: MSCs from male donor rats were treated with PDGF-BB, and a novel microbubble formulation was prepared using a thin-film hydration method. In vivo, MSCs with or without PDGF-BB pretreatment were transplanted by UTMD after inducing AMI in experimental rats. The therapeutic efficacy of PDGF-BB-primed MSCs on myocardial apoptosis, angiogenesis, cardiac function and scar repair was estimated. The effects and molecular mechanisms of PDGF-BB on MSC migration and survival were explored in vitro. RESULTS: The results showed that the biological effects of UTMD increased the local levels of stromal-derived factor-1 (SDF-1), which promoted the migration of transplanted MSCs to the ischemic region. Compared with UTMD alone, UTMD combined with PDGF-BB pretreatment significantly increased the cardiac homing of MSCs, which subsequently reduced myocardial apoptosis, promoted neovascularization and tissue repair, and increased cardiac function 30 days after MI. The vitro results demonstrated that PDGF-BB enhanced MSC migration and protected these cells from H2O2-induced apoptosis. Mechanistically, PDGF-BB pretreatment promoted MSC migration and inhibited H2O2-induced MSC apoptosis via activation of the phosphatidylinositol 3-kinase/serine-threonine kinase (PI3K/Akt) pathway. Furthermore, crosstalk between PDGF-BB and stromal-derived factor-1/chemokine receptor 4 (SDF-1/CXCR4) is involved in the PI3K/AKT signaling pathway. CONCLUSION: The present study demonstrated that UTMD combined with PDGF-BB treatment could enhance the homing ability of MSCs, thus alleviating AMI in rats. Therefore, UTMD combined with PDGF-BB pretreatment may offer exciting therapeutic opportunities for strengthening MSC therapy in ischemic diseases.

Low-intensity pulsed ultrasound (LIPUS) enhances the anti-inflammatory effects of bone marrow mesenchymal stem cells (BMSCs)-derived extracellular vesicles.

BACKGROUND: Bone marrow-derived mesenchymal stem cells (BMSCs)-derived extracellular vesicles (EVs) have shown potent anti-inflammatory function in various pathological conditions, such as osteoarthritis and neurodegenerative diseases. Since the number of EVs naturally secreted by cells is finite and they usually bear specific repertoires of bioactive molecules to perform manifold cell-cell communication, but not one particular therapeutic function as expected, their practical application is still limited. Strategies are needed to increase the production of EVs and enhance their therapeutic function. Recent studies have suggested that low-intensity pulsed ultrasound (LIPUS) is a promising non-invasive method to increase the secretion of EVs and promote their anti-inflammatory effects. However, the effect of LIPUS stimulation of BMSCs on EVs derived from the cells remains unclear. The objective of this study was to investigate whether LIPUS stimulation on BMSCs could increase the secretion of EVs and enhance their anti-inflammatory effects. METHODS: BMSCs were exposed to LIPUS (300 mW/cm2) for 15 min and EVs were isolated by ultracentrifugation. Anti-inflammatory effects of EVs were investigated on RAW264.7 cells in vitro and in the allogeneic skin transplantation model. Small RNA-seq was utilized to identify components difference in EVs with/without LIPUS irradiation. RESULTS: In this study, we found that LIPUS stimulation could lead to a 3.66-fold increase in the EVs release from BMSCs. Moreover, both in vitro and in vivo experimental results suggested that EVs secreted from LIPUS-treated BMSCs (LIPUS-EVs) possessed stronger anti-inflammatory function than EVs secreted from BMSCs without LIPUS stimulation (C-EVs). RNA-seq analysis revealed that miR-328-5p and miR-487b-3p were significantly up-regulated in LIPUS-EVs compare with C-EVs. The suppression of MAPK signaling pathway by these two up-regulated miRNAs could be the potential mechanism of strengthened anti-inflammatory effects of LIPUS-EVs. CONCLUSION: LIPUS stimulation on BMSCs could significantly increase the secretion of EVs. Moreover, EVs generated from LIPUS-treated BMSCs possessed much stronger anti-inflammatory function than C-EVs. Therefore, LIPUS could be a promising non-invasive strategy to promote the production of EVs from BMSCs and augment their anti-inflammatory effects.

Enhancing Doxorubicin Delivery in Solid Tumor by Superhydrophobic Amorphous Calcium Carbonate-Doxorubicin Silica Nanoparticles with Focused Ultrasound.

The current approach of delivering chemotherapy via pH-sensitive amorphous calcium carbonate-doxorubicin silica nanoparticles (ADS NPs) faces the challenge of insufficient drug dose due to drug instability within the bloodstream and poor tumor penetration. To overcome these long-standing obstacles, we proposed a superhydrophobic coating on the surface of the ADS NPs that could be easily modified via fluorination (ADSF NPs). The surface of fluorinated ADS NPs was further modified with a phospholipid layer to reduce aggregation and improve biocompatibility (ADSFL NPs). The contact angle and mean size of ADSFL NPs were 30.2 ± 4.4° and 353.1 ± 54.2 nm, respectively. The superhydrophobic layer generated interfacial nanobubbles on the outer shell of the NPs that reduced water-induced leakage of doxorubicin (DOX) sevenfold compared with the uncoated group and induced a cavitation effect upon ultrasound (US) sonication. Moreover, release of DOX from the ADSFL NPs could be triggered by US, and this release was further improved 1.6-fold in acidic aqueous conditions, indicating that the ADSFL NPs retained pH responsiveness. Enhanced sonography contrast and histological examination demonstrated that US could trigger cavitation activities from ADSFL NPs in vivo to induce vessel disruption and enhance the fluorescence intensity of DOX within the tumor region threefold under US imaging guidance compared with the ADSFL NPs-only group.

Noninvasive Radionuclide Molecular Imaging of the CD4-Positive T Lymphocytes in Acute Cardiac Rejection.

Heart transplantation (HT) is an effective treatment for end-stage heart disease. However, acute rejection (AR) is still the main cause of death within one year after HT. AR is an acute immune response mediated by T lymphocytes, mainly CD4+ T lymphocytes. This study innovatively develops a radiolabeled probe 99mTc-HYNIC-mAbCD4 for noninvasive visualization of CD4+ T lymphocyte infiltration and detection of AR. The 99mTc-HYNIC-mAbCD4 and its isotype control 99mTc-HYNIC-IgG were successfully prepared and characterized. The specificity and affinity of the probe in vitro were assessed by cell-binding experiments. Binding of 99mTc-HYNIC-mAbCD4 to CD4+ T lymphocytes was higher than that of the macrophages and IgG probe groups, and mAbCD4 was effective in the blockade of the binding reaction. The biodistribution data confirmed the SPECT/CT images, with significantly higher levels of 99mTc-HYNIC-mAbCD4 observed in allografts compared to allograft treatment (10 mg/kg/d Cyclosporin A subcutaneously for 5 consecutive days after surgery), isografts, or in rats which received allografts injected with 99mTc-HYNIC-IgG. Histological examination confirmed more CD4+ T lymphocyte infiltration in the allograft hearts than other groups. In summary, 99mTc-HYNIC-mAbCD4 achieved high affinity and specificity of binding to CD4+ T lymphocytes and accumulation in the transplanted heart. Radionuclide molecular imaging with 99mTc-HYNIC-mAbCD4 may be a potential diagnostic method for acute cardiac rejection.

Biomimetic PLGA Microbubbles Coated with Platelet Membranes for Early Detection of Myocardial Ischaemia-Reperfusion Injury.

Early diagnosis of myocardial ischaemia-reperfusion (MI/R) injury is important for protecting the myocardium and improving patient prognoses. Fortunately, the platelet membrane possesses the ability to target the region of MI/R injury. Therefore, we hypothesized that platelet membrane-coated particles (PMPs) could be used to detect early MI/R injury by ultrasound imaging. We designed PMPs with a porous polylactic-co-glycolic acid (PLGA) core coated with a platelet membrane shell. Red blood cell membrane-coated particles (RMPs) were fabricated as controls. Transmission electron microscopy (TEM) and fluorescence microscopy were applied to confirm the membrane coatings of the PMPs and RMPs. In vitro imaging of the PMPs and RMPs was verified. Moreover, binding experiments were designed to examine the targeting ability of the PMPs. Finally, we assessed the signal intensity of the adherent PMPs in the risk area and remote area by ultrasound imaging based on an MI/R rat model. The platelet membrane equipped the PMPs with an accurate targeting ability. Compared with RMPs, PMPs showed significantly more adhesion to human umbilical vein endothelial cells and collagen IV in vitro. Both PMPs and RMPs exhibited good enhancement ability in vitro and in vivo. Furthermore, the signal intensity of PMPs in the risk area was significantly higher than that in remote areas. These results were further validated by an immunofluorescence assay and ex vivo fluorescence imaging. In summary, ultrasound imaging with PMPs can detect early MI/R injury in a noninvasive manner.

Roles of Textural and Surface Properties of Nanoparticles in Ultrasound-Responsive Systems.

Acoustic inertial cavitation (IC) is a crucial phenomenon for many ultrasound (US)-related applications. This study aimed to investigate the roles of textural and surface properties of NPs in IC generation by combining typical IC detection methods with various types of silica model NPs. Acoustic passive cavitation detection, optical high-speed photography, and US imaging have been used to quantify IC activities (referred to as the IC dose, ICD) and describe the physical characteristics of IC activities from NPs. The results showed that the ICDs from NPs were positively correlated to their surface hydrophobicity and that their external surface hydrophobicity plays a much more crucial role than do the textural properties. The high-speed photography revealed that the sizes of IC-generated bubbles from superhydrophobic NPs ranged from 20-40 µm at 4-6 MPa and collapsed in several microseconds. Bubble clouds monitored with US imaging showed that IC from NPs was consistent with the surface hydrophobicity. The simulation results based on the crevice model of cavitation nuclei correlated well with the experimental results. This study has demonstrated that the surface property, instead of the textural property, of NPs dominated the IC generation, and surface nanobubbles adsorbed on the NP surface have been proposed to be cavitation nuclei.

Progress of Piezoelectric Semiconductor Nanomaterials in Sonodynamic Cancer Therapy.

Sonodynamic therapy is an emerging noninvasive tumor treatment method that utilizes ultrasound to stimulate sonosensitizers to produce a large amount of reactive oxygen species, inducing tumor cell death. Though sonodynamic therapy has very promising prospects in cancer treatment, the application of early organic sonosensitizers has been limited in efficacy due to the high blood clearance-rate, poor water solubility, and low stability. Inorganic sonosensitizers have thus been developed, among which piezoelectric semiconductor materials have received increasing attention in sonodynamic therapy due to their piezoelectric properties and strong stability. In this review, we summarized the designs, principles, modification strategies, and applications of several commonly used piezoelectric materials in sonodynamic therapy and prospected the future clinical applications for piezoelectric semiconductor materials in sonodynamic therapy.

A novel FK506-loading mesoporous silica nanoparticle homing to lymph nodes for transplant rejection treatment.

Tacrolimus (FK506) is an effective therapeutic for transplant rejection in clinical practice, primarily inhibiting rejection by suppressing the activation and proliferation of allogeneic T cells in the lymph nodes (LNs). However, conventional administration methods face challenges in directly delivering free FK506 to the LNs. In this study, we introduce a novel LN-targeted delivery system based on mesoporous silica nanoparticles (MSNs-FK506-MECA79). These particles were designed to selectively target high endothelial venules in LNs; this was achieved through surface modification with MECA79 antibodies. Their mean size and zeta potential were 201.18 ± 5.98 nm and - 16.12 ± 0.36 mV, respectively. Our findings showed that MSNs-FK506-MECA79 could accumulate in LNs and increase the local concentration of FK506 from 28.02 ± 7.71 ng/g to 123.81 ± 76.76 ng/g compared with the free FK506 treatment group. Subsequently, the therapeutic efficacy of MSNs-FK506-MECA79 was evaluated in a skin transplantation model. The treatment with MSNs-FK506-MECA79 could lead to a decrease in the infiltration of T cells in the grafts, a reduction in the grade of rejection, and a significant prolongation of survival. Consequently, this study presents a promising strategy for the active LN-targeted delivery of FK506 and improving the immunotherapeutic effects on transplant rejection.

Platelet membrane-derived biomimetic microbubbles with enhanced targeting ability for the early detection of myocardial ischemia-reperfusion injury.

Myocardial ischemia-reperfusion injury (MIRI) is a widely recognized cardiovascular disease that significantly impacts the prognosis of patients undergoing myocardial infarction recanalization. This condition can be fatal and involves complex pathophysiological mechanisms. Early diagnosis of MIRI is crucial to minimize myocardial damage and reducing mortality. Based on the inherent relationship between platelets and MIRI, we developed biomimetic microbubbles coated with platelet membrane (MB-pla) for early identification of MIRI. The MB-pla were prepared through a recombination process involving platelet membrane obtained from rat whole blood and phospholipids, blended in appropriate proportions. By coating the microbubbles with platelet membrane, MB-pla acquired various adhesion molecules, thereby gaining the capability to selectively adhere to damaged endothelial cells in the context of MIRI. In vitro experiments demonstrated that MB-pla exhibited remarkable targeting characteristics, particularly toward type IV collagen and human umbilical vein endothelial cells that had been injured through hypoxia/reoxygenation procedures. In a rat model of MIRI, the signal intensity produced by MB-pla was notably higher than that of control microbubbles. These findings were consistent with results obtained from fluorescence imaging of isolated hearts and immunofluorescence staining of tissue sections. In conclusion, MB-pla has great potential as a non-invasive early detection method for MIRI. Furthermore, this approach can potentially find application in other conditions involving endothelial injury in the future.

A Membrane-Targeting Aggregation-Induced Emission Probe for Monitoring Lipid Droplet Dynamics in Ischemia/Reperfusion-Induced Cardiomyocyte Ferroptosis.

Myocardial ischemia/reperfusion injury (MIRI) is the leading cause of irreversible myocardial damage. A pivotal pathogenic factor is ischemia/reperfusion (I/R)-induced cardiomyocyte ferroptosis, marked by iron overload and lipid peroxidation. However, the impact of lipid droplet (LD) changes on I/R-induced cardiomyocyte ferroptosis is unclear. In this study, an aggregation-induced emission probe, TPABTBP is developed that is used for imaging dynamic changes in LD during myocardial I/R-induced ferroptosis. TPABTBP exhibits excellent LD-specificity, superior capability for monitoring lipophagy, and remarkable photostability. Molecular dynamics (MD) simulation and super-resolution fluorescence imaging demonstrate that the TPABTBP is specifically localized to the phospholipid monolayer membrane of LDs. Imaging LDs in cardiomyocytes and myocardial tissue in model mice with MIRI reveals that the LD accumulation level increase in the early reperfusion stage (0-9 h) but decrease in the late reperfusion stage (>24 h) via lipophagy. The inhibition of LD breakdown significantly reduces the lipid peroxidation level in cardiomyocytes. Furthermore, it is demonstrated that chloroquine (CQ), an FDA-approved autophagy modulator, can inhibit ferroptosis, thereby attenuating MIRI in mice. This study describes the dynamic changes in LD during myocardial ischemia injury and suggests a potential therapeutic target for early MIRI intervention.

Ultrasound-Responsive Biomimetic Superhydrophobic Drug-Loaded Mesoporous Silica Nanoparticles for Treating Prostate Tumor.

Interfacial nanobubbles on a superhydrophobic surface can serve as ultrasound cavitation nuclei for continuously promoting sonodynamic therapy, but their poor dispersibility in blood has limited their biomedical application. In this study, we proposed ultrasound-responsive biomimetic superhydrophobic mesoporous silica nanoparticles, modified with red blood cell membrane and loaded with doxorubicin (DOX) (F-MSN-DOX@RBC), for RM-1 tumor sonodynamic therapy. Their mean size and zeta potentials were 232 ± 78.8 nm and -35.57 ± 0.74 mV, respectively. The F-MSN-DOX@RBC accumulation in a tumor was significantly higher than in the control group, and the spleen uptake of F-MSN-DOX@RBC was significantly reduced in comparison to that of the F-MSN-DOX group. Moreover, the cavitation caused by a single dose of F-MSN-DOX@RBC combined with multiple ultrasounds provided continuous sonodynamic therapy. The tumor inhibition rates in the experimental group were 71.5 8 ± 9.54%, which is significantly better than the control group. DHE and CD31 fluorescence staining was used to assess the reactive oxygen species (ROS) generated and the broken tumor vascular system induced by ultrasound. Finally, we can conclude that the combination of anti-vascular therapy, sonodynamic therapy by ROS, and chemotherapy promoted tumor treatment efficacy. The use of red blood cell membrane-modified superhydrophobic silica nanoparticles is a promising strategy in designing ultrasound-responsive nanoparticles to promote drug-release.

Ultrasonic Microbubble Cavitation Deliver Gal-3 shRNA to Inhibit Myocardial Fibrosis after Myocardial Infarction.

Galectin-3 (Gal-3) participates in myocardial fibrosis (MF) in a variety of ways. Inhibiting the expression of Gal-3 can effectively interfere with MF. This study aimed to explore the value of Gal-3 short hairpin RNA (shRNA) transfection mediated by ultrasound-targeted microbubble destruction (UTMD) in anti-myocardial fibrosis and its mechanism. A rat model of myocardial infarction (MI) was established and randomly divided into control and Gal-3 shRNA/cationic microbubbles + ultrasound (Gal-3 shRNA/CMBs + US) groups. Echocardiography measured the left ventricular ejection fraction (LVEF) weekly, and the heart was harvested to analyze fibrosis, Gal-3, and collagen expression. LVEF in the Gal-3 shRNA/CMB + US group was improved compared with the control group. On day 21, the myocardial Gal-3 expression decreased in the Gal-3 shRNA/CMBs + US group. Furthermore, the proportion of the myocardial fibrosis area in the Gal-3 shRNA/CMBs + US group was 6.9 ± 0.41% lower than in the control group. After inhibition of Gal-3, there was a downregulation in collagen production (collagen I and III), and the ratio of Col I/Col III decreased. In conclusion, UTMD-mediated Gal-3 shRNA transfection can effectively silence the expression of Gal-3 in myocardial tissue, reduce myocardial fibrosis, and protect the cardiac ejection function.

Targeted microRNA delivery by lipid nanoparticles and gas vesicle-assisted ultrasound cavitation to treat heart transplant rejection.

Despite exquisite immune response modulation, the extensive application of microRNA therapy in treating heart transplant rejection is still impeded by poor stability and low target efficiency. Here we have developed a low-intensity pulsed ultrasound (LIPUS) cavitation-assisted genetic therapy after executing the heart transplantation (LIGHT) strategy, facilitating microRNA delivery to target tissues through the LIPUS cavitation of gas vesicles (GVs), a class of air-filled protein nanostructures. We prepared antagomir-155 encapsulated liposome nanoparticles to enhance the stability. Then the murine heterotopic transplantation model was established, and antagomir-155 was delivered to murine allografted hearts via the cavitation of GVs agitated by LIPUS, which reinforced the target efficiency while guaranteeing safety owing to the specific acoustic property of GVs. This LIGHT strategy significantly depleted miR-155, upregulating the suppressors of cytokine signaling 1 (SOCS1), leading to reparative polarization of macrophages, decrease of T lymphocytes and reduction of inflammatory factors. Thereby, rejection was attenuated and the allografted heart survival was markedly prolonged. The LIGHT strategy achieves targeted delivery of microRNA with minimal invasiveness and great efficiency, paving the way towards novel ultrasound cavitation-assisted strategies of targeted genetic therapy for heart transplantation rejection.

New trends in non-pharmacological approaches for cardiovascular disease: Therapeutic ultrasound.

Significant advances in application of therapeutic ultrasound have been reported in the past decades. Therapeutic ultrasound is an emerging non-invasive stimulation technique. This approach has shown high potential for treatment of various disease including cardiovascular disease. In this review, application principle and significance of the basic parameters of therapeutic ultrasound are summarized. The effects of therapeutic ultrasound in myocardial ischemia, heart failure, myocarditis, arrhythmias, and hypertension are explored, with key focus on the underlying mechanism. Further, the limitations and challenges of ultrasound therapy on clinical translation are evaluated to promote application of the novel strategy in cardiovascular diseases.

Amplification of Plasma MicroRNAs for Non-invasive Early Detection of Acute Rejection after Heart Transplantation With Ultrasound-Targeted Microbubble Destruction.

OBJECTIVE: Acute rejection (AR) screening has always been the focus of patient management in the first several years after heart transplantation (HT). As potential biomarkers for the non-invasive diagnosis of AR, microRNAs (miRNAs) are limited by their low abundance and complex origin. Ultrasound-targeted microbubble destruction (UTMD) technique could temporarily alter vascular permeability through cavitation. We hypothesized that increasing the permeability of myocardial vessels might enhance the abundance of circulating AR-related miRNAs, thus enabling the non-invasive monitoring of AR. METHODS: The Evans blue assay was applied to determine efficient UTMD parameters. Blood biochemistry and echocardiographic indicators were used to ensure the safety of the UTMD. AR of the HT model was constructed using Brown-Norway and Lewis rats. Grafted hearts were sonicated with UTMD on postoperative day (POD) 3. The polymerase chain reaction was used to identify upregulated miRNA biomarkers in graft tissues and their relative amounts in the blood. RESULTS: Amounts of six kinds of plasma miRNA, including miR-142-3p, miR-181a-5p, miR-326-3p, miR-182, miR-155-5p and miR-223-3p, were 10.89 ± 1.36, 13.54 ± 2.15, 9.84 ± 0.70, 8.55 ± 2.00, 12.50 ± 3.96 and 11.02 ± 3.47 times higher in the UTMD group than those in the control group on POD 3. Plasma miRNA abundance in the allograft group without UTMD did not differ from that in the isograft group on POD 3. After FK506 treatment, no miRNAs increased in the plasma after UTMD. CONCLUSION: UTMD can promote the transfer of AR-related miRNAs from grafted heart tissue to the blood, allowing non-invasive early detection of AR.

Electrospun polymer fibers modified with FK506 for the long-term treatment of acute cardiac allograft rejection in a heart transplantation model.

FK506, a first-line immunosuppressant, is routinely administered orally and intravenously following heart transplantation. However, frequent administration can result in a substantial psychological burden to patients, resulting in non-adherence to medication. The purpose of our study is to overcome the disadvantages of systemic drug administration by developing a polymer-based delivery system that is tunable and biodegradable and that can release highly hydrophobic FK506 over extended periods to treat or prevent acute cardiac allograft rejection. Using an electrospinning method, long-acting microfibers were prepared, and FK506 appeared to be continuously released for up to 14 days based on the in vitro release profiles. After implanting the microfiber subcutaneously into the abdominals of transplanted rats, it was found that the infiltration of T cells and macrophages and the secretion of interleukin-2 (IL-2) and IL-1ß were significantly reduced compared with those of the free FK506 groups. More importantly, the mean survival time (MST) of the PCL-FK506 group was significantly extended in comparison with that of untreated control recipients and free FK506 (MST of untreated control recipients, free FK506, and PCL-FK506 was 8, 26.1, and 37, respectively). In conclusion, we propose that this drug delivery approach would be suitable for developing long-lasting immunomodulatory agents that prolong cardiac graft survival safely and effectively.

Granzyme B-responsive fluorescent probe for non-invasive early diagnosis of transplant rejection.

Allograft rejection has always been a major obstacle in organ transplantation. The current clinical diagnostic gold standard for allograft rejection is an invasive biopsy. However, biopsy has some limitations, such as sampling errors, risk of serious complications, and high cost. In this study, we have rationally developed an activatable fluorescent probe CYGB for imaging of granzyme B, which is a biomarker released by CD8+T cells attacking the graft. Moreover, the ability of CYGB to detect rejection early in mouse heart and skin transplantation models was evaluated. The probe CYGB consists of a caged hemicyanine-based fluorophore and a GzmB-specifically cleaved peptide substrate linked via a self-immolating spacer, p-aminobenzyl alcohol. Endogenous GzmB in CD8+ T cells specifically activated the near-infrared fluorescence (NIRF) signal of CYGB. In vivo imaging in mice skin and heart graft models, showed that CYGB preferentially accumulates in grafts, enabling early diagnosis of rejection. Moreover, CYGB enables non-invasive assessment of the level of immunosuppression in allogeneic mice treated with FK506. This study provides an alternative method for monitoring the status of allografts without biopsy.