Adhesion Of Cells Research Articles

Abstract 2823: Development of an in vitro blood-brain barrier model of CCR7/CCL19 induced invasion of the central nervous system

Abstract Pediatric T-cell acute lymphoblastic leukemia (T-ALL), an aggressive hematological malignancy, is characterized by abnormal thymocyte proliferation, frequently with mutations of the Notch-1 gene and has an overall 10-year survival rate of ~27%. A marker of poor prognosis is the presence of T-ALL within the central nervous system (CNS), which is treated with intrathecal chemotherapy and cranial irradiation; two toxic treatments with high morbidities. Gene-expression profiling of Notch-1-induced adhesion regulators showed that C-C chemokine receptor 7 (CCR7) was highly expressed and was associated with leukemic T-cell invasion of the CNS. Normal functions of CCR7 include lymph node homing and immune cell trafficking in response to CCR7 ligands, CC-chemokine ligand 19 (CCL19) or CCL21. In several studies CCL19 has been linked to T-cell invasion of the CNS across the blood-brain barrier (BBB). We hypothesize that antagonizing CCR7 binding to CCL19 will block migration of T-ALL into the CNS and thereby alleviate the need for toxic therapies. To test our hypothesis, we are using receptor internalization assays, calcium mobilization assays, transwell migration assays and an in vitro model of an intact human BBB. We have used flow cytometry-based CCR7 internalization assays ± CCR7 antagonist and found that we can completely block CCL19-induced receptor internalization in human CCR7-expressing CCRF-CEM, and HuT-78 T-ALL cells, and chemotaxis via β1 integrins across a fibronectin-coated transwell membrane. Our BBB employs HBEC-5i, human cerebral microvascular endothelial cells, in transwell chambers coated with gelatin and fibronectin for the adhesion of endothelial cells. HBEC-5i cells are a suitable cell line to make a 3D model of the BBB, as they form an endothelial monolayer with tight junctions (TJ), which is crucial to maintaining the integrity and inhibiting permeability of the BBB. We are establishing a transwell migration assay to assess the migration of CCRF-CEM T-ALL cells across the HBEC-5i cell monolayer. The integrity of the HBEC-5i cell monolayer is assessed by trans-epithelial electrical resistance (TEER), which measures the electrical resistance across the HBEC-5i monolayer, a measure of TJ integrity. Using TEER measurements, we observe that HBEC-5i form a monolayer that reaches an intact monolayer functioning with TJs over 72-96 hours. We are using this model system to investigate the migration of CCR7-expressing CCRF-CEM cells through the BBB ± CCL19. Subsequently, we will add the CCL19 antagonist (CCL197-77) to determine if inhibiting CCR7 is sufficient to eliminate migration of the CCRF-CEM cells through the BBB. If successful, this model will allow us to test the contribution of CCR7 signaling to integrins, the contribution of other immune cell types and the overall molecular mechanisms that mediate invasion of the CNS in a 3-D model of the BBB. Citation Format: Alondra Rodriguez, Cesar I. Cardona, Colin A. Bill, Charlotte M. Vines. Development of an in vitro blood-brain barrier model of CCR7/CCL19 induced invasion of the central nervous system [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2823.

Abstract 119: A non-surgical method for developing a pre-clinical orthotopic mouse model for colorectal cancer

Abstract Recapitulating physiologically relevant tumor microenvironment is important yet a significant challenge in modeling colorectal cancer (CRC) for basic and translational studies. Among the different methods, using the cecal-wall injection technique is the most common way to create animal models of colorectal cancer (CRC). However, this approach requires complex surgery, and its unnatural placement creates a significant obstacle for developing treatments. Here, we report a non-surgical procedure for establishing orthotopic CRC mouse models, reflecting the appropriate tumor microenvironment to better model the disease development and facilitate translational studies including drug identification and validation. In this procedure, anesthesia is induced in overnight fasted study immunocompetent or immunodeficient mice while placed on a thermostat-controlled heating blanket. A specialized catheter is inserted and distended into the mouse colon to gently block the colon and ensure the localization for tumor development. Hereafter, the colonic epithelium is treated with proteases and mechanical abrasion to cause mild disruption. Following the local gut inflammation, the targeted colon region is incubated with tumor cells, leading to the adhesion of tumor cells to the colonic epithelium. Based on the animal background and the cell line used, the tumor developed within 3-4 weeks. The stages of tumor development at different time points were successfully confirmed using hematoxylin and eosin stain. Tumor microenvironment data revealed a significant increase in the immune cell infiltration specifically CD4+ T-cells and tumor-associated macrophages in the non-surgical procedure as compared to the cecal-wall injection. Furthermore, a significant increase in the PD-1 expression of the CD8+ T-cells in the cecal-wall injection as compared to the non-surgical procedure, suggests that the precise location of the tumor site may exert a substantial influence on pre-clinical therapeutic outcomes of immune-checkpoint inhibitors, thereby potentially hampering the clinical efficacy of drug treatments. Additional adaptations of this procedure may include: i) tumor development in different mouse strains, ii) local delivery of adeno-Cre in transgenic mice for CRC, and iii) delivering tumor organoids and patient-derived xenografts in immuno-deficient mice. Overall, our non-surgical procedure overcomes the technical difficulties of surgery and offers several advantages like ease of handling, cost-effectiveness, pain-free with nearly no procedure-associated mortality and adaptability to several strains. In conclusion, this procedure provides novel and promising alternative to conventional surgical orthotopic CRC mouse models for therapeutic interventions. Citation Format: Naresh Singh, Samantha Sharma, Kevin Van Der Jeught, Zhuolong Zhou, Xinna Zhang, Xiongbin Lu. A non-surgical method for developing a pre-clinical orthotopic mouse model for colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 119.

Abstract 5410: Unveiling PAI-1 as a new player in chemoresistant melanoma in aged human skin reconstructed in vitro

Abstract The impact of advanced glycation end products (AGEs) accumulation is associated with the development and exacerbation of many degenerative processes and disorders, including aging and cancer. AGEs accumulation in the skin leads to stiffness and loss of elasticity by building up in the connective tissue during aging process. This process has implications of the extracellular matrix, affecting the adhesion, migration, and invasiveness potential of melanoma cells. Furthermore, tumor invasiveness is correlated with mechanical stiffness, contributing to the high metastatic capacity of melanoma. To address this research gap, our study aimed to evaluate the invasive potential of chemoresistant melanoma using an in vitro model of reconstructed aged human skin modified by glycation. Glycated skin exhibited distinct features indicative of aging, including positive pentosidine fluorescence and elevated CML labeling and expression, well-established biomarkers of aging. Furthermore, glycated skin with resistant melanoma demonstrated reduced expression of differentiated keratinocytes (CK-10) and increased expression of undifferentiated keratinocytes (CK-14). Overall, in glycated skin with resistant melanoma, decreased proliferation and increased tumor invasion were observed, associated with changes in the expression of key markers, such as reduced MITF and SOX-10 and increased AXL. Downregulation of MITF in skin with resistant melanoma resulted in decreased Melan-A expression, implying altered pigmentation of melanoma cells. Additionally, resistant melanoma in glycated skin showed enhanced inflammation, as indicated by higher levels of IL-8 and IL-1α. Gene screening showed upregulation of the PAI-1, ABL-1, COL1A1, SPARC genes. Among these genes, PAI-1 (Plasminogen activator inhibitor-1) was upregulated in resistant melanoma compared to naïve melanoma both in glycated skin. PAI-1 may play a critical role in the development of aging-associated pathological changes. In addition, PAI-1 is recognized as a marker of senescence and a key member of a group of proteins collectively known as the senescence-messaging secretome. It also performs protumorigenic functions, such as promoting vasculature, increasing growth, motility and protecting tumor cell apoptosis. Overall, this work demonstrated that skin aged by glycation when associated with i-Braf resistant melanoma exhibited a more invasive and less proliferative profile (low MITF and SOX10, elevated AXL), more inflammatory (elevated IL-8 and IL-1α) and with a greater number of immature keratinocytes (elevated ck-14). PAI-1 overexpression may be a promising target to expand studies caused by oncogenes in the model of aged skin by glycation with resistant melanoma. Citation Format: Renaira Oliveira da Silva, Larissa A. Carvalho, Denisse E. Camarena, Julia R. Silva, Isabella H. Noma, Manoel O. Moraes, Paula C. Pennacchi, Silvya S. Maria Engler. Unveiling PAI-1 as a new player in chemoresistant melanoma in aged human skin reconstructed in vitro [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5410.

Porous Titanium/SF Sponge Composite Scaffold with Loads of Gentamicin and Small Extracellular Vesicles to Improve Antibacterial and Osteogenesis Properties

AbstractAdditive manufacturing has been widely used for producing porous titanium orthopedic implants. However, the higher surface area inherent to porous structures increases the risk of bacterial colonization. In this work, composite scaffolds with dual functions of the antibacterial and osteogenesis promotion properties are developed. The porous titanium is fabricated using a selective laser melting process to provide mechanical support. Silk fibrin (SF) sponge‐loading gentamicin and extracellular vesicles (EVs) are prepared in the pores of the porous titanium. The developed composite scaffolds are investigated for their antibacterial and osteogenesis effects. The samples exhibit well antibacterial activity against Escherichia coli. The samples also show good cytocompatibility, effectively promoting the adhesion and proliferation of the bone marrow mesenchymal stem cells (rBMSCs). EVs significantly promote the expression of osteogenesis‐related genes in rBMCSs and the superior extracellular matrix mineralization reveals that EVs have a remarkable role in enhancing osteogenic differentiation. Therefore, the composite scaffolds with dual functions of antibacterial and osteogenesis promotion properties have excellent potential for orthopedic implant applications.

Fabrication of 3D porous polyurethane-graphene oxide scaffolds by a sequential two-step processing for non-load bearing bone defects

Bone defects as a common orthopedic disease lead to severe pains over a long period. Scaffolds are novel approaches in tissue engineering to treat bone problems and deal with their challenges. Here, 3D porous polyurethane (PU) scaffolds containing graphene oxide (GO) with different percentages (0, 0.1, 0.3, and 0.5 wt%) were developed through a combination of freeze-drying and salt etching techniques for bone tissue engineering applications. The morphologies of scaffolds, physicochemical properties, the degree of crystallinity, and hydrophilicity were evaluated by SEM, FTIR, XRD, and water contact angle assay, respectively. The porosity, degradation behavior, compressive strength, and elastic modulus of 3D porous scaffolds were also determined. To assess the scaffold bioactivity, the morphology of the deposited calcium phosphate layer on the scaffold with macro-structure was evaluated by SEM images. The viability and adhesion of MG63 osteoblast-like cells cultured on the fabricated scaffolds were examined by MTT assay and SEM images, respectively. The results show that adding GO particles not only had no effect on the interconnectivity and porosity of 3D porous macroscopic structures of neat PU but also smaller and more uniformed microscopically pores were obtained. The crystallinity, water contact angle, and weight loss of scaffolds increased as the higher GO concentrations were employed. Followed by increasing GO contents from 0 to 0.5 wt%, the compressive strength and Young’s modulus were increased by 232% and 245%, respectively. The bioactivity of scaffolds was fostered as GO concentration increased. Although, the MTT assay proved the biocompatibility of PU scaffolds containing 0.1 and 0.3 wt% GO, the samples loaded with 0.5 GO had a negative impact on the viability of MG63 cell lines. In conclusion, the present study demonstrates a high potential of PU scaffolds loaded with 0.1 and 0.3 wt% GO particles in bone tissue engineering applications.

Functionalization of 3D printed Ti6Al4V high-porous spinal implant surface with use of plasma electrolytic oxidation

Highly-porous titanium implants manufactured using additive methods open new perspectives for reconstructive medicine. Ti6Al4V alloy is one of the most frequently used biomaterials in medicine. It has proven biocompatibility and methods of modifying it to improve functionality have been extensively investigated. However, the translation of the current state of knowledge to products manufactured using additive methods is not clear. Thecomplex architecture of scaffolds creates many challenges and limitations for surface modification, especially in internal parts of implants. The paper presents a developed surface modification using plasma electrolytic oxidation to improve the physicochemical properties of a spinal implant intended for the treatment of cervical discopathy. The properties of the produced surface layer were assessed based on SEM observations with EDS analysis, optical profilometry, XPS and XRD analysis, corrosion test and plasma atomic emission spectroscopy. The proposed modifications had a positive impact on the corrosion resistance of implants, limited the penetration of metal ions into the environment imitating the tissue environment, and significantly influenced the surface topographies, which should constitute a better substrate for the adhesion and proliferation of bone cells.

Listeria monocytogenes: the silent assassin.

Listeriosis is a foodborne infection in humans caused by Listeria monocytogenes. Consumption of contaminated food can lead to severe infection in vulnerable patients, that can be fatal. Clinical manifestations include sepsis and meningitis, and in pregnancy-associated infection, miscarriage and stillbirth. Diagnosis is confirmed by culture and identification of the pathogen from blood, cerebrospinal fluid, vaginal swab, placenta or amniotic fluid. Treatment regimens recommend amoxicillin, ampicillin or an aminoglycoside. Virulence factors mediate bacterial adhesion and invasion of gut epithelial cells. Other factors mediate biofilm formation and tolerance to low temperatures and high salt concentrations facilitating persistence and survival in the environment.

Innovative hydroxyapatite/submicron mesoporous SiO2/HA particles composite coatings for enhanced osteogenic activity of NiTi bone implants: A comprehensive investigation of materials and biological interactions

In this study, hydroxyapatite (HA) incorporated in mesoporous SiO2 particles (SiO2/HA) was initially synthesized using a soft template method. The resulting structure owned a high surface area (796.62 m2 g-1) and a substantial pore volume (0.757 cm3 g-1). The TEM and FESEM images indicated that the synthesized mesoporous particles possessed a spherical morphology, with a diameter of approximately 200 nm, and pores of slit-shape. The mission was to improve the surface characteristics of NiTi implants by composite coatings. This investigation emphasized specifically the impact of different concentrations of mesoporous SiO2/HA particles within the electrolyte on the properties of HA/mesoporous composite coatings. The surface characteristics of the coatings were examined using ATR-FTIR, FESEM, EDS, AFM, and hydrophilicity measurements. Furthermore, the osteogenic activity of the coatings was evaluated by culturing the osteosarcoma cell line SAOS-2 on the coated samples. The introduction of mesoporous particles resulted in a decrease in the coatings’ surface roughness along with an improvement in their wettability. Moreover, there was an enhancement in the cell viability (99% after 7 days), proliferation, adhesion, and growth of SAOS-2 cells at the optimized concentration of mesoporous SiO2/HA particles (1000 mg/L). DAPI images illustrated that the mesoporous particles did not cause any adverse impact on the cellular nuclei. Finally, in the apoptosis test, the composite coating displayed a reduced number of necrotic cells compared to the pure HA coating.

Scratch resistance and in-vitro biocompatibility of plasma-sprayed baghdadite coatings reinforced with carbon nanotubes

Hydroxyapatite (HA, Ca10(PO4)6(OH)2) coatings, commonly used for metallic implants, have limitations such as excessive brittleness, low fracture toughness, inadequate wear resistance, and slow osseointegration properties. In contrast, baghdadite (BAG, Ca3ZrSi2O9), a calcium zirconium silicate-based bioceramic, exhibits outstanding biological properties, to promote cell proliferation and differentiation of human osteoblasts along with adequate mechanical strength. In this study, plasma-sprayed HA, BAG, and carbon nanotubes (CNT, 1 wt%, and 2 wt%) reinforced BAG coatings are deposited on titanium (Ti) substrate to enhance its mechanical as well as biological properties. The microhardness values of CNT-reinforced BAG coatings increase due to a decrease in porosity and the retention of CNT inside the BAG matrix. Progressive loading scratch tests are performed, revealing a reduction in wear volume loss from 15.051 mm3 to 12.574 mm3 and scratch rate from 43.262 mm3N−1 m−1 to 36.172 mm3N−1 m−1 with the addition of 2 wt% CNT in BAG coatings. The scratch hardness test results demonstrated that introducing CNT into BAG coating significantly enhances its performance, showing a remarkable 38.7 % improvement. In-vitro cell culture studies indicate excellent cell adhesion, growth, and proliferation of MC3T3-E1 osteoblast cells on the surfaces of the CNT-reinforced BAG coatings.

Retraction: Insulin-like Growth Factor-1 Induces Adhesion and Migration in Human Multiple Myeloma Cells via Activation of β1-Integrin and Phosphatidylinositol 3'-Kinase/AKT Signaling.

Retraction: Insulin-like Growth Factor-1 Induces Adhesion and Migration in Human Multiple Myeloma Cells via Activation of β1-Integrin and Phosphatidylinositol 3'-Kinase/AKT Signaling.

Titanium dioxide bioceramics prepared by 3D printing method and its structure effect on stem cell behavior

3D printing was widely used for the modification of bioceramic structures. Here, Bioactive titanium dioxide (TiO2) ceramics with macropore structures were prepared by controlling the pore size through 3D printing, and further obtained secondary micropores by altering the sintering temperature. The structures and in vitro bioactivity of materials were characterized, and the properties of the ceramics for osteogenic differentiation of rabbits bone marrow mesenchymal stem cells (rBMSCs) were investigated. The results showed that titanium dioxide ceramics could induce apatite formation in SBF, which indicated it is bioactive. All materials exhibited porous structures, which could promote the proliferation and osteogenic differentiation of rBMSCs. Importantly, the structure with 200 μm pore size significantly promoted the proliferation and adhesion of cells. Furthermore, more secondary micropores formed on the surface of materials with the decreased sintering temperature, which promoted cell proliferation. The pore size of 400 μm significantly upregulated the expression of osteogenesis-related genes, including Col I, ALP, Runx2, and OPN. It had the strongest osteogenic ability when sintered at 1100 °C compared to other groups. This implies that the 3D-printed TiO2 ceramics have good osteogenic properties, and the hierarchical structure consisting of macroscopic pores and secondary micropores can control it well.

REDV-Functionalized Recombinant Spider Silk for Next-Generation Coronary Artery Stent Coatings: Hemocompatible, Drug-Eluting, and Endothelial Cell-Specific Materials.

Coronary artery stents are life-saving devices, and millions of these devices are implanted annually to treat coronary heart disease. The current gold standard in treatment is drug-eluting stents, which are coated with a biodegradable polymer layer that elutes antiproliferative drugs to prevent restenosis due to neointimal hyperplasia. Stenting is commonly paired with systemic antiplatelet therapy to prevent stent thrombosis. Despite their clinical success, current stents have significant limitations including inducing local inflammation that drives hyperplasia; a lack of hemocompatibility that promotes thrombosis, increasing need for antiplatelet therapy; and limited endothelialization, which is a critical step in the healing process. In this research, we designed a novel material for use as a next-generation coating for drug-eluting stents that addresses the limitations described above. Specifically, we developed a recombinant spider silk material that is functionalized with an REDV cell-adhesive ligand, a peptide motif that promotes specific adhesion of endothelial cells in the cardiovascular environment. We illustrated that this REDV-modified spider silk variant [eADF4(C16)-REDV] is an endothelial-cell-specific material that can promote the formation of a near-confluent endothelium. We additionally performed hemocompatibility assays using human whole blood and demonstrated that spider silk materials exhibit excellent hemocompatibility under both static and flow conditions. Furthermore, we showed that the material displayed slow enzyme-mediated degradation. Finally, we illustrated the ability to load and release the clinically relevant drug everolimus from recombinant spider silk coatings in a quantity and at a rate similar to that of commercial devices. These results support the use of REDV-functionalized recombinant spider silk as a coating for drug-eluting stents.

Zeaxanthin impairs angiogenesis and tumor growth of glioblastoma: An in vitro and in vivo study

ObjectivesTo investigate the therapeutic effects of Zeaxanthin (Zea), one of the oxidized xanthophyll carotenoids belonging to the isoprenoids, on inhibiting the angiogenesis and tumor growth of glioblastoma (GBM) via an in vitro and in vivo study. MethodsThe effects of Zea on the proliferation, adhesion, migration and invasion of human GBM cell lines were detected by cell proliferation assay, cell adhesion assay and Transwell assay. The effect of Zea on angiogenesis was detected by rat aortic ring assay and human umbilical vein endothelial cells (HUVEC) in vitro tube formation assay. The effects of Zea on PARP, Caspase 3 and VEGFR2 phosphorylation as well as VEGFR2's downstream signaling pathway were detected by Western blot. The in vivo human GBM xenograft mouse model was employed to study the therapeutic efficacy of Zea. ResultsZea impaired the proliferation, adhesion, migration and invasion of U87 and U251 cells as well as HUVECs. Rat aortic ring experiments displayed Zea significantly inhibited angiogenesis during VEGF-induced microvascular germination. In vitro and in vivo vascular experiments verified that Zea inhibited VEGF-induced HUVEC proliferation and capillary-like tube formation. Additionally, Zea induced GBM cells apoptosis via increasing the expression of cleaved PARP and Caspase 3. In HUVECs and U251 GBM cells, Zea down-regulated VEGF-induced activation of the VEGFR2 kinase pathway. Meanwhile the expression of p-AKT, p-ERK, p-STAT3 and FAK were all attenuated in U251 cells. Moreover, the effects of Zea on GBM cells proliferation could be blocked by VEGFR2 kinase inhibitor SU5408. These results suggest that Zea may hinder GBM angiogenesis and tumor growth through down-regulating a cascade of oncogenic signaling pathways, both through the inhibition of angiogenesis and the anti-tumor mechanism of a direct cytotoxic effect. Besides, Zea inhibits GBM angiogenesis and tumor growth exemplified through a xenograft mouse model in vivo. ConclusionZea impairs angiogenesis and tumor growth of GBM both in vitro and in vivo. It can be declared that Zea is a potential valuable anticancer candidate for the future treatment strategy of GBM.

Microbiologic profile of Escherichia coli strains isolated from urinary tract infections in aged nephrology clinic patients.

The invasion of bacteria in the urinary system in humans characterizes the urinary tract infection (UTI) with the capacity to cause cystitis and/or pyelonephritis. Escherichia coli is the main microorganism associated with UTI and was therefore called UPEC (Uropathogenic Escherichia coli). UTI prevalence between genders decreases after 65 years in community infections and in post-menopausal women. Urinary infections can occur in outpatients or inpatients in the presence of several factors such as differentiated antimicrobial resistance profile, attachment, and invasion to uroepithelial cells, presence of virulence factors, biofilm formation, and prevalent phylogenetic group. The present study aims to relate the UTI by UPEC in outpatients older than 55 years attended in nephrology. It was detected that virulence mechanisms such as adhesion and invasion in renal cells may be associated with the urinary tract infection process caused by UPEC in these patients. Biofilm formation and association with phylogenetic group D can be important determinants of early adhesion and infection.

Staphylococcal superantigen-like protein 3 triggers murine mast cell adhesion by binding to CD43 and augments mast cell activation.

Staphylococcus aureus is a noteworthy pathogen in allergic diseases, as four staphylococcal exotoxins activate mast cells, a significant contributor to inflammation, in an IgE-independent manner. Although the adhesion of mast cells is an essential process for their immune responses, only a small number of exotoxins have been reported to affect the process. Here, we demonstrated that staphylococcal superantigen-like (SSL) 3, previously identified as a toll-like receptor 2 agonist, induced the adhesion of murine bone marrow-derived mast cells to culture substratum. SSL3-induced adhesion was mediated by fibronectin in an Arg-Gly-Asp (RGD) sequence-dependent manner, suggesting the integrins were involved in the process. Additionally, SSL3 was found to bind to an anti-adhesive surface protein CD43. SSL3 induced the adhesion of HEK293 cells expressing exogenous CD43, suggesting that CD43 is the target molecule for adhesion induced by SSL3. Evaluation of SSL3-derived mutants showed that the C-terminal region (253-326), specifically T285 and H307, are necessary to induce adhesion. SSL3 augmented the IL-13 production of mast cells in response to immunocomplex and SSL12. These findings reveal a novel function of SSL3, triggering cell adhesion and enhancing mast cell activation. This study would clarify the correlation between S. aureus and allergic diseases such as atopic dermatitis.

The dual HCK/BTK inhibitor KIN-8194 impairs growth and integrin-mediated adhesion of BTKi-resistant mantle cell lymphoma

Although Bruton’s tyrosine kinase (BTK) inhibitors (BTKi) have significantly improved patient prognosis, mantle cell lymphoma (MCL) is still considered incurable due to primary and acquired resistance. We have recently shown that aberrant expression of the Src-family tyrosine kinase hematopoietic cell kinase (HCK) in MCL correlates with poor prognosis, and that genetic HCK perturbation impairs growth and integrin-mediated adhesion of MCL cells. Here, we show that KIN-8194, a dual inhibitor of BTK and HCK with in vivo activity against Myd88-L265P-driven diffuse large B-cell lymphoma and Waldenström Macroglobulinemia, has a potent growth inhibitory effect in MCL cell lines and primary MCL cells, irrespective of their sensitivity to BTKi (ibrutinib and acalabrutinib). In BTKi-resistant cells this is mediated by inhibition of HCK, which results in repression of AKT-S6 signaling. In addition, KIN-8194 inhibits integrin-mediated adhesion of BTKi-sensitive and insensitive MCL cells to fibronectin and stromal cells in an HCK-dependent manner. Finally, we show that MCL cells with acquired BTKi resistance retain their sensitivity to KIN-8194. Taken together, our data demonstrate that KIN-8194 inhibits growth and integrin-mediated adhesion of BTKi-sensitive MCL cells, as well as MCL cells with primary or acquired BTKi resistance. This renders KIN-8194 a promising novel treatment for MCL patients.

The anticancer activity of bovine lactoferrin is reduced by deglycosylation and it follows a different pathway in cervix and colon cancer cells.

Bovine lactoferrin (bLF) is a glycosylated protein with purported beneficial properties. The aim of this work was to determine the role of bLF glycosylation in the adhesion, internalization, and growth inhibition of cancer cells. The viability of cervix (HeLa) and colon (Caco-2) cancer cells (MTT assay and epifluorescence microscopy) was inhibited by bLF, while deglycosylated bLF (bLFdeg) had no effect. Adhesion to cell surfaces was quantified by immunofluorescence assay and showed that bLF was able to bind more efficiently to both cell lines than bLFdeg. Microscopic observations indicated that bLF glycosylation favored bLF binding to epithelial cells and that it was endocytosed through caveolin-1-mediated internalization. In addition, the mechanism of action of bLF on cancer cell proliferation was investigated by determining the amount of phosphorylated intermediates of signaling pathways such as epidermal growth factor receptor (EGFR), extracellular signal-regulated kinase (ERK) and protein kinase B (known as Akt). Chemoluminescence immunoassay of phosphorylated intermediates showed that bLF inhibited Akt phosphorylation, consistent with its growth inhibiting activity. This assay also indicated that the bLF receptor/signaling pathways may be different in the two cell lines, Caco-2 and HeLa. This work confirmed the effect of glycosylated bLF in inhibiting cancer cell growth and that glycosylation is required for optimal surface adhesion, internalization, and inhibition of the ERK/Akt pathway of cell proliferation through glycosylated cell surface receptors.

A Multifunctional Metal-Phenolic Nanocoating on Bone Implants for Enhanced Osseointegration via Early Immunomodulation.

Surface modification is an important approach to improve osseointegration of the endosseous implants, however it is still desirable to develop a facile yet efficient coating strategy. Herein, a metal-phenolic network (MPN) is proposed as a multifunctional nanocoating on titanium (Ti) implants for enhanced osseointegration through early immunomodulation. With tannic acid (TA) and Sr2+ self-assembled on Ti substrates, the MPN coatings provided a bioactive interface, which can facilitate the initial adhesion and recruitment of bone marrow mesenchymal stem cells (BMSCs) and polarize macrophage toward M2 phenotype. Furthermore, the TA-Sr coatings accelerated the osteogenic differentiation of BMSCs. In vivo evaluations further confirmed the enhanced osseointegration of TA-Sr modified implants via generating a favorable osteoimmune microenvironment. In general, these results suggest that TA-Sr MPN nanocoating is a promising strategy for achieving better and faster osseointegration of bone implants, which can be easily utilized in future clinical applications.

Self-Assembled DNA Composite-Engineered Mesenchymal Stem Cells for Improved Skin-Wound Repair.

The direct use of mesenchymal stem cells (MSCs) as therapeutics for skin injuries is a promising approach, yet it still faces several obstacles, including limited adhesion, retention, and engraftment of stem cells in the wound area, as well as impaired regenerative and healing functions. Here, DNA-based self-assembled composites are reported that can aid the adhesion of MSCs in skin wounds, enhance MSC viability, and accelerate wound closure and re-epithelialization. Rolling-circle amplification (RCA)-derived DNA flowers, equipped with multiple copies of cyclic Arg-Gly-Asp (cRGD) peptides and anti-von Willebrand factor (vWF) aptamers, act as robust scavengers of reactive oxygen species (ROS) and enable synergistic recognition and adhesion to stem cells and damaged vascular endothelial cells. These DNA structure-aided stem cells are retained at localized wound sites, maintain repair function, and promote angiogenesis and growth factor secretion. In both normal and diabetes-prone db/db mice models with excisional skin injuries, facile topical administration of DNA flower-MSCs elicits rapid blood vessel formation and enhances the sealing of the wound edges in a single dose. DNA composite-engineered stem cells warrant further exploration as a new strategy for the treatment of skin and tissue damage.

Synergistic Effect of B-Type Lotus Seedpod Oligomeric Procyanidin and Probiotics against Adhesion of Enterotoxigenic Escherichia coli In Vitro

Enterotoxigenic Escherichia coli (ETEC) adhesion to gut epithelial cells is a prerequisite for diarrhea. Here, we studied the synergistic effect and potential mechanism of B-type lotus seedpod oligomeric procyanidin (LSPC) combined with probiotics against adhesion of ETEC. The results indicated that LSPC exhibited an effective anti-ETEC adhesion effect. LSPC showed signally synergistic effects with probiotics, e.g., Streptococcus thermophilus (ST) and Lactobacillus rhamnosus (LGG) in response to ETEC adhesion, with the combination of LSPC and LGG being the most efficacious. This may be attributed to the restoration of transmembrane resistance of cells, the increased expression of anti-inflammatory factors (IL-10, 1.89-fold), and the reduction of cellular inflammatory factor levels (TNF-α and IL-8), regulated by LGG-LSPC, resulting in a better enhancement of cellular immune defense and barrier function. In addition, the results of gas chromatography showed that LSPC, as a prebiotic, could significantly increase the total amount of short-chain fatty acids (especially, butyric acid) produced by probiotics (e.g., LGG), thus better maintaining intestinal health against ETEC infection. In conclusion, the synergistic effect of LSPC and probiotics (represented by LGG) against ETEC adhesion in epithelial cells may be achieved through the enhancement of cellular immune defense, cellular barrier function, and maintenance of homeostasis in the gut.