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Sérgio Scalzo,1 Anderson K Santos,1 Heloísa AS Ferreira,1 Pedro A Costa,1 Pedro HDM Prazeres,2 Natália JA da Silva,1 Lays C Guimarães,1 Mário de Morais e Silva,1 Marco TR Rodrigues Alves,1 Celso TR Viana,1 Itamar CG Jesus,1 Alice P Rodrigues,1 Alexander Birbrair,2 Anderson O Lobo,3 Frederic Frezard,1 Michael J Mitchell,4 Silvia Guatimosim,1 Pedro Pires Goulart Guimaraes1 1Department of Physiology and Biophysics, Federal University of Minas Gerais, Belo Horizonte, MG, Brazil; 2Department of General Pathology, Federal University of Minas Gerais, Belo Horizonte, MG, Brazil; 3Department of Materials Engineering, Federal University of Piauí, Teresina, PI, Brazil; 4Department of Bioengineering, University of Pennsylvania, Philadelphia, PA, USACorrespondence: Pedro Pires Goulart Guimaraes, Department of Physiology and Biophysics, Federal University of Minas Gerais, Belo Horizonte, MG, Brazil, Tel +55 031 3409-2948, Email ppiresgo@reitoria.ufmg.brIntroduction: Gene therapy is a promising approach to be applied in cardiac regeneration after myocardial infarction and gene correction for inherited cardiomyopathies. However, cardiomyocytes are crucial cell types that are considered hard-to-transfect. The entrapment of nucleic acids in non-viral vectors, such as lipid nanoparticles (LNPs), is an attractive approach for safe and effective delivery.Methods: Here, a mini-library of engineered LNPs was developed for pDNA delivery in cardiomyocytes. LNPs were characterized and screened for pDNA delivery in cardiomyocytes and identified a lead LNP formulation with enhanced transfection efficiency.Results: By varying lipid molar ratios, the LNP formulation was optimized to deliver pDNA in cardiomyocytes with enhanced gene expression in vitro and in vivo, with negligible toxicity. In vitro, our lead LNP was able to reach a gene expression greater than 80%. The in vivo treatment with lead LNPs induced a twofold increase in GFP expression in heart tissue compared to control. In addition, levels of circulating myeloid cells and inflammatory cytokines remained without significant changes in the heart after LNP treatment. It was also demonstrated that cardiac cell function was not affected after LNP treatment.Conclusion: Collectively, our results highlight the potential of LNPs as an efficient delivery vector for pDNA to cardiomyocytes. This study suggests that LNPs hold promise to improve gene therapy for treatment of cardiovascular disease.Graphical Abstract: Keywords: lipid nanoparticles, ionizable lipids, pDNA delivery, heart, cardiomyocytes

Beyond its actions on the nervous system, amitriptyline (AM) has been shown to lower inflammatory, angiogenic, and fibrogenic markers in a few pathological conditions in human and in experimental animal models. However, its effects on foreign body reaction (FBR), a complex adverse healing process, after biomedical material implantation are not known. We have evaluated the effects of AM on the angiogenic and fibrogenic components on a model of implant-induced FBR. Sponge disks were implanted subcutaneously in C57BL/6 mice, that were treated daily with oral administration of AM (5 mg/kg) for seven consecutive days in two protocols: treatment was started on the day of surgery and the implants were removed on the seventh day after implantation and treatment started 7 days after implantation and the implants removed 14 after implantation. None of the angiogenic (vessels, Vascular endothelial growth factor (VEGF), and interleukin-1β (IL-1β) or fibrogenic parameters (collagen, TGF-β, and fibrous capsule) and giant cell numbers analyzed were attenuated by AM in 7-day-old implants. However, AM was able to downregulate angiogenesis and FBR in 14-day-old implants. The effects of AM described here expands its range of actions as a potential agent capable of attenuating fibroproliferative processes that may impair functionality of implantable devices., (© 2023 International Union of Biochemistry and Molecular Biology.)

The use of implantable biomaterials to replace physiological and anatomical functions has been widely investigated in the clinic. However, the selection of biomaterials is crucial for long-term function, and the implantation of certain biomaterials can cause inflammatory and fibrotic processes, triggering a foreign body reaction that leads to loss of function and consequent need for removal. Specifically, the Wnt signaling pathway controls the healing process of the human body, and its dysregulation can result in inflammation and fibrosis, such as in peritoneal fibrosis. Here, we assessed the effects of daily oral administration of a Wnt pathway inhibitor complex (CD:LGK974) to reduce the inflammatory, fibrotic, and angiogenic processes caused by intraperitoneal implants. CD:LGK974 significantly reduced the infiltration of immune cells and release of inflammatory cytokines in the implant region compared to the control groups. Furthermore, CD:LGK974 inhibited collagen deposition and reduced the expression of pro-fibrotic α-SMA and TGF-β1, confirming fibrosis reduction. Finally, the CD:LGK974 complex decreased VEGF levels and both the number and area of blood vessels formed, suggesting decreased angiogenesis. This work introduces a potential new application of the Wnt inhibitor complex to reduce peritoneal fibrosis and the rejection of implants at the intraperitoneal site, possibly allowing for longer-term functionality of existing clinical biomaterials., (© 2023. Controlled Release Society.)

New Findings: What is the central question of this study? Peritoneal injury can result in a persistent fibroproliferative process in the abdominal cavity, causing pain and loss of function of internal organs. This study aimed to demonstrate the use of sodium butyrate (NaBu) as a potential agent to attenuate peritoneal fibrosis induced by a synthetic matrix. What is the main finding and its importance? Our findings provide the first evidence that NaBu attenuates the inflammatory, angiogenesis and fibrogenesis axes involved in the formation of peritoneal fibrovascular tissue, indicating the potential of this compound to ameliorate peritoneal fibrosis., Abstract: The aim of this study was to identify the bio-efficacy of sodium butyrate (NaBu) on preventing the development of peritoneal fibrovascular tissue induced by implantation of a synthetic matrix in the abdominal cavity. Polyether-polyurethane sponge discs were implanted in the peritoneal cavity of mice, which were treated daily with oral administration of NaBu (100 mg/kg). Control animals received water (100 μl). After 7 days, the implants were removed for assessment of inflammatory, angiogenic and fibrogenic markers. Compared with control values, NaBu treatment decreased mast cell recruitment/activation, inflammatory enzyme activities, levels of pro-inflammatory cytokines, and the proteins p65 and p50 of the nuclear factor-κB pathway. Angiogenesis, as determined by haemoglobin content, vascular endothelial growth factor levels and the number of blood vessels in the implant, was reduced by the treatment. In NaBu-treated animals, the predominant collagen present in the abdominal fibrovascular tissue was thin collagen, whereas in control implants it was thick collagen. Transforming growth factor-β1 levels were also lower in implants of treated animals. Sodium butyrate downregulated the inflammatory, angiogenesis and fibrogenesis axes of the fibroproliferative tissue induced by the intraperitoneal synthetic matrix. This compound has potential to control/regulate chronic inflammation and adverse healing processes in the abdominal cavity., (© 2022 The Authors. Experimental Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Implantation of biomedical/synthetic devices to replace and/or repair biological tissues very often induces an adverse healing response (scarce angiogenesis, excessive collagen deposition) which is detrimental to implant functionality and integration to host tissue. Interleukin-33/ST2 axis (IL-33/ST2) has been shown to modulate angiogenic and remodeling processes in several types of injuries. However, its effects on these processes after implantation of synthetic matrix have not been reported. Using synthetic matrix of polyether-polyurethane implanted subcutaneously in mice lacking ST2 receptor (ST2/KO), we characterized neovascularization and matrix remodeling in the fibrovascular tissue induced by the implants. Tissue accumulation was increased inside and around the implants in KO implants relative to the wild type (WT). More intense proliferative activity, using CDC 47 marker, was observed in KO implants compared with that of WT implants. Angiogenesis, using two endothelial cell markers, Von Willebrand Factor (VWF) and vascular endothelial cell VE cadherin and hemoglobin content, increased in implants of KO mice relative to control WT. Remodeling of the newly formed fibrovascular tissue (soluble collagen and PicroSirius Red-stained histological sections) showed predominance of type 1 collagen in ST2-KO implants versus type 3 in control implants. The number of positive cells for caspase-3, apoptotic marker, decreased in ST2 group. Our findings evidenced a role of IL-33/ST2 axis in restraining blood vessel formation and regulating the pattern of matrix remodeling in the fibrovascular tissue induced by synthetic implants. Intervention in this cytokine complex holds potential to accelerate integration of biomaterial and host tissue by improving blood supply and matrix remodeling., (Copyright © 2021. Published by Elsevier Inc.)

Background: Colorectal cancer (CRC) is the third most common cancer in the world. 5- Fluorouracil (5-FU) is a conventional and most effective drug used in the clinic for the treatment of CRC. However, the clinical use of 5-FU is limited due to the acquired resistance and systemic toxicity, such as hepatotoxicity and gastrointestinal toxicity., Objective: Recent advances in nanomedicine are being exploited to develop nanoparticle platforms to overcome resistance and therapeutic delivery of active molecules. Here, we developed 5-FU loaded sulfadiazine-poly(lactide-co-glycolide) nanoparticles (SUL-PLGA NPs) to be applied in the colorectal cancer model., Methods: We assessed the in vivo efficacy of the SUL-PLGA NPs to enhance the antitumor effect of 5-FU., Results: In vivo treatment with 5-FU-SUL-PLGA NPs significantly reduced tumor growth in a colon cancer xenograft model compared to free 5-FU and 5-FU loaded non-targeted NPs. Treatment with 5-FU-SUL-PLGA NPs also increased blood vessel diameters within tumors, which could act in conjunction to enhance antitumor efficacy. In addition, 5-FU-SUL-PLGA NPs significantly reduced liver mass and lung mass, which are the most common metastasis sites of CRC, and decreased liver hepatotoxicity compared to free 5-FU drug and 5-FU loaded non-targeted NPs., Conclusion: Our findings suggest that the use of 5-FU-SUL-PLGA NPs is a promising strategy to enhance 5-FU efficacy against CRC., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

A number of genetic factors have been linked to the development of diabetes, a condition that often requires implantable devices such as glucose sensors. In normoglycaemic individuals, this procedure induces a foreign body reaction (FBR) that is detrimental to bioimplant functionality. However, the influence of the genetic background on this reaction in diabetes has not been investigated. We examined the components of FBR (capsule thickness, collagen deposition, mast cell and foreign body giant cell number) in subcutaneous implants of polyether polyurethane (SIPP) in streptozotocin (STZ)-induced diabetes in Swiss, C57BL/6 and Balb/c mice. The fasting blood glucose levels before STZ injections were 133.5 ± 5.1 mg/dL, after the treatment increased 68.4% in Swiss mice, 62.4% in C57BL/6 and 30.9% in Balb/c mice. All FBR features were higher in implants of Swiss and C57BL/6 mice compared with those in implants of Balb/c. Likewise, the apoptotic index was higher in implants of diabetic Swiss and C57BL/6 mice whose glycaemic levels were the highest. Our findings show an association between the severity of hyperglycaemic levels and the intensity of the FBR to SIPP. These important strain-related differences in susceptibility to diabetes and the intensity of the FBR must be considered in management using implantable devices in diabetic individuals., (© 2021 Company of the International Journal of Experimental Pathology (CIJEP).)

Recent data has signaled that in addition to its therapeutic indications as antidepressant and analgesic, amitriptyline (AM) exerts anti-inflammatory effects in humans and experimental animal models of acute inflammation. We tested the hypothesis that this compound could also modulate the chronic inflammatory process induced by synthetic matrix in mice. Polyether-polyurethane sponge disks were implanted subcutaneously in 9-week-old male C57BL/6 mice. The animals received by oral gavage 5.0 mg/kg of amitriptyline for seven consecutive days in two treatment regimens. In the first series, the treatment was initiated on the day of surgery and the implants removed at day 7 post-implantation. For the assessment of the effect of amitriptyline on chronic inflammation, the treatment was initiated 7 days post-implantation and the sponge discs removed 14 after implantation. The inflammatory markers evaluated, myeloperoxidase - MPO, nitrite content, IL-6, IFN-γ, TNF-α, CXCL1 and CCL2 levels, and NF-κB transcription factor activation were reduced in implants when the treatment began 7 days post-implantation (chronic inflammation). In contrast, only mast cell number, MPO activity and activation of NF-κB pathway decreased when the treatment began soon after implantation (sub-acute inflammation) in 7-day old implants. The anti-inflammatory effects of amitriptyline described here, extend its range of actions as a potential agent able to attenuate long-term inflammatory processes.

The damaging effects of obesity extend to multiple pre-existing tissue/organs. However, the influence of this condition on key components (inflammation and angiogenesis) of fibrovascular connective proliferating tissue, essential in repair processes, has been neglected. Our objective in this study was to investigate whether obesity would influence inflammatory-angiogenesis induced by synthetic matrix of polyether-polyurethane implanted subcutaneously in high-fat-fed obese C57/BL6 mice. Fourteen days after implantation, the inflammatory and angiogenic components of the newly formed tissue intra-implant were evaluated. The pro-inflammatory enzyme activities, myeloperoxidase (MPO) and N-acetyl-β-D-glucosaminidase (NAG), the levels of TNF-α, CXCL1/KC and CCL2 and NF-κB transcription factor were examined. Angiogenesis was determined by morphometric analysis of implant blood vessels, intra-implant levels of hemoglobin content, VEGF levels, and western blot for VEGFR2. All inflammatory and angiogenic markers were increased in the implants of obese mice compared with their non-obese counterparts. Similarly, activation of the NF-κB pathway and phosphorylation of VEGFR2 were higher in implants of obese mice (1.60 ± 0.28 Np65/Cp65; 0.96 ± 0.08 p-VEGFR2/VEGFR2-T) compared with implants of non-obese animals (1.40 ± 0.14; 0.49 ± 0.08). These observations suggest that obesity exerts critical role in sponge-induced inflammatory-angiogenesis, possibly by activating fibrovascular components in the inflamed microenvironment. Thus, this pathological condition causes damage not only to pre-existing tissues/organs but also to newly formed proliferating fibrovascular tissue. This is relevant to the development of therapeutic approaches to improve healing processes in patients with obesity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

IL-33/ST2 axis has been shown to exert both pro- and anti- effects in wound healing and tumor development. To further understand the role of this cytokine complex, we characterized comparatively the inflammatory component of a malignant tissue and non-malignant tissue in mice lacking ST2 receptor (ST2-KO). KO mice and their wild-type (WT) counterparts were either implanted subcutaneously with polyether-polyurethane sponge discs to induce non-malignant fibrovascular tissue growth or inoculated with 4T1 cells to induce mammary tumor. Loss of ST2 receptor in mice resulted in enhanced mammary tumor and fibrovascular tissue relative to the WT animals. The inflammatory parameters (MPO and NAG activities, levels of the cytokines CXCL1/KC, CCL2, TNF-α, TGF-β1, and mast cell number) were differentially modulated in both tissues. In tumors, these parameters were, overall, lower compared with those in tumors of WT mice. In KO implants, CXCL1/KC and TNF-α levels increased; MPO, NAG, and CCL2 levels decreased relative to the WT implants. In addition, deletion of ST2 receptor inhibited mast cell recruitment but had no effect on TGF-β1 levels in implants. Our study has shown antitumorigenic effect of ST2 in mammary tumor and this may be mediated by downregulation of pro-inflammatory cytokines (CXCL1/KC, CCL2, TNF-α, and TGF-β1). Conversely, in the fibrovascular tissue, lack of ST2 receptor resulted in differential modulation of cytokine production. Differential signaling mechanisms may be activated by IL-33/ST2 axis to modulate cytokine production in malignant and non-malignant proliferative processes.

Activation of the Wnt signaling pathway promotes lung cancer progression and contributes to poor patient prognosis. The porcupine inhibitor LGK974, a novel orally bioavailable cancer therapeutic in Phase I clinical trials, induces potent Wnt signaling inhibition and leads to suppressed growth and progression of multiple types of cancers. The clinical use of LGK974, however, is limited in part due to its low solubility and high toxicity in tissues that rely on Wnt signaling for normal homeostasis. Here, we report the use of host-guest chemistry to enhance the solubility and bioavailability of LGK974 in mice through complexation with cyclodextrins (CD). We assessed the effects of these complexes to inhibit Wnt signaling in lung adenocarcinomas that are typically driven by overactive Wnt signaling. 2D 1 H NMR confirmed host-guest complexation of CDs with LGK974. CD:LGK974 complexes significantly decreased the expression of Wnt target genes in lung cancer organoids and in lung cancer allografts in mice. Further, CD:LGK974 complexes increased the bioavailability upon oral administration in mice compared to free LGK974. In a mouse lung cancer allograft model, CD:LGK974 complexes induced potent Wnt signaling inhibition with reduced intestinal toxicity compared to treatment with free drug. Collectively, the development of these complexes enables safer and repeated oral or parenteral administration of Wnt signaling inhibitors, which hold promise for the treatment of multiple types of malignancies., (Copyright © 2018. Published by Elsevier B.V.)

Background/objectives: Pancreas regenerative capacity after injury is not always sufficient to comply with the body's requirement of digestive enzymes and hormones. We present an alternative system to induce pancreas parenchyma proliferation (exocrine and endocrine components), rather than regeneration or remodeling in normoglycemic mice., Methods: Porous discs of polyether-polyurethane were surgically placed adjacent to the native pancreas and removed at days 15, 30 and 45 after implantation. No exogenous growth factors or extracellular matrix components were added to the platform. The synthetic matrix provided a platform that was filled with parenchymal and non-parenchymal pancreas tissue as detected by histological analysis. Immunohistochemistry analysis were performed to identify insulin positive cells in the newly formed tissue. In addition, angiogenic, inflammatory and metabolic parameters were carried out in those mice., Results: At day 15, the pores of the platform were filled with inflammatory cells, spindled-shaped like fibroblasts, extracellular matrix components, blood vessels and clusters of pancreatic parenchyma (acini, ducts and islet-like structures). At days 30 and 45 the pancreas features remained well organized; its organization resembled that of a native pancreas. Interestingly, besides islet-like structures that showed positive cells to insulin, some ductal cells were also positive for insulin immunostaining. No significant differences in serum glucose and c-peptide concentrations during the experimental period were detected., Conclusions: The plain synthetic porous platform (without addition of exogenous molecules) placed adjacent to the native organ exhibits potential to restore and/or expand exocrine (acini, ducts) and endocrine (β-cell mass) components in pancreatic injuries and in high metabolic demand., (Copyright © 2017 IAP and EPC. Published by Elsevier B.V. All rights reserved.)

Objective: Obesity is a highly prevalent multifactorial metabolic condition in which the need for functional bioengineered substitutes (e.g., scaffolds for tissue engineering) is likely to occur. However, the adverse foreign body response (FBR) that invariably takes place adjacent to implant devices impairing their function is poorly characterized in this condition. This study investigated the influence of obesity on the host response to a synthetic matrix implanted subcutaneously in high-fat-fed obese mice., Methods: Histological analysis of 14-day-old implants was performed to identify collagen deposition, capsule thickness, fibroblast-like cells, foreign body giant cells, and mast cells. In addition, transforming growth factor β1 (TGF-β1) levels in the implants and serum were determined., Results: All fibrogenic markers (and TGF-β1 levels) increased in the implants of obese mice compared with their nonobese counterparts. Particularly relevant was the fibrous capsule thickness in implants of obese mice (234.2 ± 22.1 µm vs. 109.2 ± 13.4 µm in implants of nonobese animals)., Conclusions: The study results showing that obesity upregulates the main features of the FBR induced by subcutaneous implants in mice may be relevant in understanding biomaterial integration and performance in this condition. This is crucial to the development of strategies to maintain the integrity and function of implantable devices., (© 2018 The Obesity Society.)

Aims: Currently, animal models of liver regeneration are based on extensive lesions of the native organ and on cellular approaches using biomaterials to host growth factors and extracellular components to create artificial liver systems. We report a polymeric biological platform, minimally invasive, that induced sequential proliferation of liver parenchyma inside the scaffold in mice., Main Methods: Porous discs of polyether-polyurethane were surgically placed under the left liver lobe and removed at days 4, 8, 12 and 25 after implantation. No exogenous growth factors or extracellular matrix components were added to the scaffold. Histological analysis of the implants was performed to identify hepatocytes, liver vascular structures and bile ducts in the newly formed tissue. In addition, systemic markers for hepatic function were determined., Key Findings: This biohybrid device provided a scaffold that was gradually filled with parenchymal and non-parenchymal liver tissue as detected by histological analysis. At day 4, the pores of the scaffold were filled with inflammatory cells and spindled-shaped like fibroblasts, and extracellular matrix components. At day 8, hepatocytes clusters, central lobular hepatic veins, portal space containing arteries, veins and biliary ducts were detected. By days 12 and 25 a liver-like structure filled 2/3 of the scaffold. Its organization resembled that of a mature liver. Serum concentration of ALT increased three-fold initially after implantation, returning gradually to control levels., Significance: The plain synthetic scaffold (without addition of exogenous molecules) placed under the intact left liver lobe exhibits the potential to investigate physiological mechanisms that regulate liver parenchyma proliferation., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Aims: Several alternative cellular approaches using biomaterials to host insulin-producing cells derived from stem cells have been developed to overcome the limitations of type 1 diabetes treatment (exogenous insulin injection). However, none seem to fulfill all requirements needed to induce pancreatic cells successful colonization of the scaffolds. Here, we report a polymeric platform adherent to the native mice pancreas filled with human adipose stem cells (hASCs) that was able to induce growth of pancreatic parenchyma., Main Methods: Synthetic polyether-polyurethane discs were placed adjacent to pancreas of normoglycemic and streptozotocin-induced diabetic mice. At day 4 post implantation, 1×10 6 hASCs were injected intra-implant in groups of normoglycemic and diabetic mice. Immunohistochemistry analysis of the implants was performed to identify insulin positive cells in the newly formed tissue. In addition, metabolic, inflammatory and angiogenic parameters were carried out in those mice., Key Findings: This study provides evidence of the ability of a biohybrid device to induce the growth of differentiated pancreas parenchyma in both normoglycemic and streptozotocin-induced diabetic mice as detected by histological analysis. Glucose metabolism and body weight of hyperglycemic mice bearing hASCs implants improved., Significance: The synthetic porous scaffold bearing hASC cells placed adjacent to the native animal pancreas exhibits the potential to be exploited in future cell-based type 1 diabetes therapies., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Roots of Smilax species (Smilacaceae), named as salsaparrilha, have been used for centuries in Asia and Americas as depurative (=for "cleaning blood"), diuretic and sudorific. In southeast of Brazil, roots of Herreria salsaparrilha Mart. (Agavaceae) are also named as salsaparrilha and are used for the same purpose. In this study, we have evaluated the antihyperlipidemic and antihyperglycemic effects of extracts from roots of Smilax brasiliensis and H. salsaparrilha in mice fed with high-refined carbohydrate diet (HC). The chemical composition of the products was determined by LC-DAD and LC-MS. Groups of mice that received the HC diet showed an increase in plasma concentrations of glucose, triglycerides and total cholesterol, compared to control group, without treatment (p<0.05). Triglycerides were reduced significantly (p<0.05) in HC diet group that received 100 and 200mg/kg BW/day of both salsaparrilha extracts. Glucose and total cholesterol levels were reduced significantly (p<0.05) in the groups that received the higher doses (200mg/kg BW/day) of both extracts of salsaparrilha. Extracts of S. brasiliensis, at this dose also showed a higher reduction in triglycerides levels (p<0.001) and promoted a significant reduction in the adipocyte area (p<0.05). Both extracts showed the presence of saponins in LC analysis but S. brasiliensis has a higher concentration of phenolics, mainly chlorogenic acid. The presence of steroidal saponins might be responsible for the reduction of the cholesterol levels, while phenolics in S. brasiliensis by the metabolism of triglycerides and better fat distribution. The result is according with the traditional use of these plants and shown their potential for use as functional foods., (Copyright © 2015 Elsevier Ltd. All rights reserved.)