Lung cancer immunotherapy: progress, pitfalls, and promises.

Lung cancer is the primary cause of mortality in the United States and around the globe. Therapeutic options for lung cancer treatment include surgery, radiation therapy, chemotherapy, and targeted drug therapy. Medical management is often associated with the development of treatment resistance leading to relapse. Immunotherapy is profoundly altering the approach to cancer treatment owing to its tolerable safety profile, sustained therapeutic response due to immunological memory generation, and effectiveness across a broad patient population. Different tumor-specific vaccination strategies are gaining ground in the treatment of lung cancer. Recent advances in adoptive cell therapy (CAR T, TCR, TIL), the associated clinical trials on lung cancer, and associated hurdles are discussed in this review. Recent trials on lung cancer patients (without a targetable oncogenic driver alteration) reveal significant and sustained responses when treated with programmed death-1/programmed death-ligand 1 (PD-1/PD-L1) checkpoint blockade immunotherapies. Accumulating evidence indicates that a loss of effective anti-tumor immunity is associated with lung tumor evolution. Therapeutic cancer vaccines combined with immune checkpoint inhibitors (ICI) can achieve better therapeutic effects. To this end, the present article encompasses a detailed overview of the recent developments in the immunotherapeutic landscape in targeting small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC). Additionally, the review also explores the implication of nanomedicine in lung cancer immunotherapy as well as the combinatorial application of traditional therapy along with immunotherapy regimens. Finally, ongoing clinical trials, significant obstacles, and the future outlook of this treatment strategy are also highlighted to boost further research in the field.

Gelatin-polycaprolactone-nanohydroxyapatite electrospun nanocomposite scaffold for bone tissue engineering.

Bone injuries and fractures generally take a long period to heal itself. To address this problem, bone tissue engineering (BTE) has gained significant research impetus. Among the several techniques used for scaffold fabrication, electrospinning ought to be the most promising technique for the development of the nanostructured scaffolds. The present study was carried out to fabricate an electrospun nanocomposite scaffold for BTE by using gelatin, polycaprolactone (PCL), and nanohydroxyapatite (nHAp). To prepare Gelatin-PCL-nHAp nanocomposite scaffold: Gelatin-PCL blend was electrospun and then treated with nHAp (1 wt%) for different time periods. The fabricated nanocomposite scaffold was analysed by field emission scanning electron microscopy (FESEM) to determine the fiber diameter and evaluate the fiber morphology. The Gelatin-PCL-nHAp nanocomposite scaffold-20 min exhibited the average fiber diameter of 615±269 nm and average pore size 4.7±1.04 µm, and also revealed the presence of nHAp particles over the Gelatin-PCL scaffold surface. Further, X-ray diffraction (XRD), Fourier Transform Infrared (FTIR) spectroscopy and thermogravimetric (TG) analysis also indicated the deposition of nHAp over the Gelatin-PCL scaffold surface. MTT assay and DNA quantification showed good viability and significant proliferation of human osteoblasts on Gelatin-PCL-nHAp nanocomposite scaffold. Moreover, cell-scaffold constructs illustrated efficient cellular attachment and adequately spread cells, and it also depicts characteristic polygonal morphology of osteoblasts over the Gelatin-PCL-nHAp nanocomposite scaffold. Thus, the results of in-vitro analysis of electrospun nanocomposite scaffold suggest that the Gelatin-PCL-nHAp scaffold can be a potential candidate for BTE applications.

Fabrication and characterization of novel nano-biocomposite scaffold of chitosan-gelatin-alginate-hydroxyapatite for bone tissue engineering.

A novel nano-biocomposite scaffold was fabricated in bead form by applying simple foaming method, using a combination of natural polymers-chitosan, gelatin, alginate and a bioceramic-nano-hydroxyapatite (nHAp). This approach of combining nHAp with natural polymers to fabricate the composite scaffold, can provide good mechanical strength and biological property mimicking natural bone. Environmental scanning electron microscopy (ESEM) images of the nano-biocomposite scaffold revealed the presence of interconnected pores, mostly spread over the whole surface of the scaffold. The nHAp particulates have covered the surface of the composite matrix and made the surface of the scaffold rougher. The scaffold has a porosity of 82% with a mean pore size of 112±19.0µm. Swelling and degradation studies of the scaffold showed that the scaffold possesses excellent properties of hydrophilicity and biodegradability. Short term mechanical testing of the scaffold does not reveal any rupturing after agitation under physiological conditions, which is an indicative of good mechanical stability of the scaffold. In vitro cell culture studies by seeding osteoblast cells over the composite scaffold showed good cell viability, proliferation rate, adhesion and maintenance of osteoblastic phenotype as indicated by MTT assay, ESEM of cell-scaffold construct, histological staining and gene expression studies, respectively. Thus, it could be stated that the nano-biocomposite scaffold of chitosan-gelatin-alginate-nHAp has the paramount importance for applications in bone tissue-engineering in future regenerative therapies.

Human dental pulp stem cells: Applications in future regenerative medicine.

Stem cells are pluripotent cells, having a property of differentiating into various types of cells of human body. Several studies have developed mesenchymal stem cells (MSCs) from various human tissues, peripheral blood and body fluids. These cells are then characterized by cellular and molecular markers to understand their specific phenotypes. Dental pulp stem cells (DPSCs) are having a MSCs phenotype and they are differentiated into neuron, cardiomyocytes, chondrocytes, osteoblasts, liver cells and ß cells of islet of pancreas. Thus, DPSCs have shown great potentiality to use in regenerative medicine for treatment of various human diseases including dental related problems. These cells can also be developed into induced pluripotent stem cells by incorporation of pluripotency markers and use for regenerative therapies of various diseases. The DPSCs are derived from various dental tissues such as human exfoliated deciduous teeth, apical papilla, periodontal ligament and dental follicle tissue. This review will overview the information about isolation, cellular and molecular characterization and differentiation of DPSCs into various types of human cells and thus these cells have important applications in regenerative therapies for various diseases. This review will be most useful for postgraduate dental students as well as scientists working in the field of oral pathology and oral medicine.

Surface modification of nanofibrous polycaprolactone/gelatin composite scaffold by collagen type I grafting for skin tissue engineering.

In the present study, a tri-polymer polycaprolactone (PCL)/gelatin/collagen type I composite nanofibrous scaffold has been fabricated by electrospinning for skin tissue engineering and wound healing applications. Firstly, PCL/gelatin nanofibrous scaffold was fabricated by electrospinning using a low cost solvent mixture [chloroform/methanol for PCL and acetic acid (80% v/v) for gelatin], and then the nanofibrous PCL/gelatin scaffold was modified by collagen type I (0.2-1.5wt.%) grafting. Morphology of the collagen type I-modified PCL/gelatin composite scaffold that was analyzed by field emission scanning electron microscopy (FE-SEM), showed that the fiber diameter was increased and pore size was decreased by increasing the concentration of collagen type I. Fourier transform infrared (FT-IR) spectroscopy and thermogravimetric (TG) analysis indicated the surface modification of PCL/gelatin scaffold by collagen type I immobilization on the surface of the scaffold. MTT assay demonstrated the viability and high proliferation rate of L929 mouse fibroblast cells on the collagen type I-modified composite scaffold. FE-SEM analysis of cell-scaffold construct illustrated the cell adhesion of L929 mouse fibroblasts on the surface of scaffold. Characteristic cell morphology of L929 was also observed on the nanofiber mesh of the collagen type I-modified scaffold. Above results suggest that the collagen type I-modified PCL/gelatin scaffold was successful in maintaining characteristic shape of fibroblasts, besides good cell proliferation. Therefore, the fibroblast seeded PCL/gelatin/collagen type I composite nanofibrous scaffold might be a potential candidate for wound healing and skin tissue engineering applications.

Fabrication and characterization of scaffold from cadaver goat-lung tissue for skin tissue engineering applications.

The present study aims to fabricate scaffold from cadaver goat-lung tissue and evaluate it for skin tissue engineering applications. Decellularized goat-lung scaffold was fabricated by removing cells from cadaver goat-lung tissue enzymatically, to have cell-free 3D-architecture of natural extracellular matrix. DNA quantification assay and Hematoxylin and eosin staining confirmed the absence of cellular material in the decellularized lung-tissue. SEM analysis of decellularized scaffold shows the intrinsic porous structure of lung tissue with well-preserved pore-to-pore interconnectivity. FTIR analysis confirmed non-denaturation and well maintainance of collagenous protein structure of decellularized scaffold. MTT assay, SEM analysis and H&E staining of human skin-derived Mesenchymal Stem cell, seeded over the decellularized scaffold, confirms stem cell attachment, viability, biocompatibility and proliferation over the decellularized scaffold. Expression of Keratin18 gene, along with CD105, CD73 and CD44, by human skin-derived Mesenchymal Stem cells over decellularized scaffold signifies that the cells are viable, proliferating and migrating, and have maintained their critical cellular functions in the presence of scaffold. Thus, overall study proves the applicability of the goat-lung tissue derived decellularized scaffold for skin tissue engineering applications.

Modification of decellularized goat-lung scaffold with chitosan/nanohydroxyapatite composite for bone tissue engineering applications.

Decellularized goat-lung scaffold was fabricated by removing cells from cadaver goat-lung tissue, and the scaffold was modified with chitosan/nanohydroxyapatite composite for the purpose of bone tissue engineering applications. MTT assay with osteoblasts, seeded over the chitosan/nanohydroxyapatite-modified decellularized scaffold, demonstrated significantly higher cell growth as compared to the decellularized scaffold without modification. SEM analysis of cell-seeded scaffold, after incubation for 7 days, represented a good cell adhesion, and the cells spread over the chitosan/nanohydroxyapatite-modified decellularized scaffold. Expression of bone-tissue-specific osteocalcin gene in the osteoblast cells grown over the chitosan/nanohydroxyapatite-modified decellularized scaffold clearly signifies that the cells maintained their osteoblastic phenotype with the chitosan/nanohydroxyapatite-modified decellularized scaffold. Therefore, it can be concluded that the decellularized goat-lung scaffold-modified with chitosan/nanohydroxyapatite composite, may provide enhanced osteogenic potential when used as a scaffold for bone tissue engineering.

Isolation of Oct4+, Nanog+ and SOX2- mesenchymal cells from peripheral blood of a diabetes mellitus patient.

Diabetes mellitus is a chronic metabolic disorder that affects millions of people worldwide. The most common form is type 2 diabetes mellitus, which results in impaired beta cell function combined with insulin resistance in peripheral organs. One recently proposed treatment approach is the use of adult stem cells derived from bone marrow in autologous stem cell transplantation. Alternatively, peripheral blood can be obtained in a more non-invasive manner. In this study, we isolated and cultured mesenchymal cells (MCs) from the peripheral blood of a diabetes mellitus patient. The cultured cells were large and elongated and had an in vitro migratory capacity in the culture dish. They expressed embryonic stem cell pluripotency markers Nanog and Oct 4 as well as mesenchymal markers CD105 and CD13, and they lacked expression of hematopoietic marker CD45. These characteristics suggest that these cells have a mesenchymal phenotype similar to that obtained from bone marrow cells. The SOX2 gene was downregulated in both the peripheral blood cells and the isolated mesenchymal cell line, indicating a defective mechanism of SOX2 in diabetes mellitus. The overall results of study demonstrate that peripheral blood can be used as a source of MCs from diabetes mellitus patients for use in future regenerative stem cell therapy and that this particular model system may be useful to study the mechanism of diabetes mellitus involving downregulation of the SOX2 cascade.

Ascorbic acid induces in vitro proliferation of human subcutaneous adipose tissue derived mesenchymal stem cells with upregulation of embryonic stem cell pluripotency markers Oct4 and SOX 2.

Mesenchymal stem cells (MSCs) have immense therapeutic potential because of their ability to self-renew and differentiate into various connective tissue lineages. The in vitro proliferation and expansion of these cells is necessary for their use in stem cell therapy. Recently our group has developed and characterized mesenchymal stem cells from subcutaneous and visceral adipose tissue. We observed that these cells show a slower growth rate at higher passages and therefore decided to develop a supplemented medium, which will induce proliferation. Choi et al. have recently shown that the use of ascorbic acid enhances the proliferation of bone marrow derived MSCs. We therefore studied the effect of ascorbic acid on the proliferation of MSCs and characterized their phenotypes using stem cell specific molecular markers. It was observed that the use of 250 µM ascorbic acid promoted the significant growth of MSCs without loss of phenotype and differentiation potential. There was no considerable change in gene expression of cell surface markers CD105, CD13, Nanog, leukemia inhibitory factor (LIF) and Keratin 18. Moreover, the MSCs maintained in the medium supplemented with ascorbic acid for a period of 4 weeks showed increase in pluripotency markers Oct4 and SOX 2. Also cells in the experimental group retained the typical spindle shaped morphology. Thus, this study emphasizes the development of suitable growth medium for expansion of MSCs and maintenance of their undifferentiated state for further therapeutic use.

Immunological effects and membrane interactions of chitosan nanoparticles.

The objective of this study was to investigate the in vitro and in vivo effects of blank chitosan nanoparticles on various molecular markers such as nitric oxide (NO) production, IL-6 gene expression, and lymphocyte proliferation involved in the wound healing process. In addition, the membrane effects of chitosan nanoparticles were evaluated using phospholipid vesicles as a model membrane. Peripheral blood mononuclear cells (PBMC) were treated with blank chitosan nanoparticles, and the effect on NO production, IL-6 gene expression, and lymphocyte proliferation was evaluated. It was observed that IL-6 gene expression was not induced at any of the doses used; however, a statistically significant dose-dependent increase in NO production was observed at doses above 68.18 microg/mL equivalent to chitosan. Furthermore, chitosan nanoparticles showed a statistically significant and dose-dependent lymphocyte proliferation as compared to the control (P < 0.05). It was observed that blank chitosan nanoparticles resulted in strong membrane perturbation when evaluated by differential scanning calorimetry studies. The in vivo effects of the blank chitosan nanoparticles were evaluated using a wound healing model. Blank chitosan nanoparticles showed significantly higher NO production in vivo as compared to the control. Overall, the study clearly indicates the immunoactivating nature of chitosan nanoparticles and their strong membrane interactive potential.