Targeting PD-L1 in cholangiocarcinoma using nanovesicle-based immunotherapy.

This study demonstrates the potential of using biological nanoparticles to deliver RNA therapeutics targeting programmed death-ligand 1 (PD-L1) as a treatment strategy for cholangiocarcinoma (CCA). RNA therapeutics offer prospects for intracellular immune modulation, but effective clinical translation requires appropriate delivery strategies. Milk-derived nanovesicles were decorated with epithelial cellular adhesion molecule (EpCAM) aptamers and used to deliver PD-L1 small interfering RNA (siRNA) or Cas9 ribonucleoproteins directly to CCA cells. In vitro, nanovesicle treatments reduced PD-L1 expression in CCA cells while increasing degranulation, cytokine release, and tumor cell cytotoxicity when tumor cells were co-cultured with T cells or natural killer cells. Similarly, immunomodulation was observed in multicellular spheroids that mimicked the tumor microenvironment. Combining targeted therapeutic vesicles loaded with siRNA to PD-L1 with gemcitabine effectively reduced tumor burden in an immunocompetent mouse CCA model compared with controls. This proof-of-concept study demonstrates the potential of engineered targeted nanovesicle platforms for delivering therapeutic RNA cargoes to tumors, as well as their use in generating effective targeted immunomodulatory therapies for difficult-to-treat cancers such as CCA.

Mesenchymal Stem Cell-Derived Exosomes Loaded with miR-155 Inhibitor Ameliorate Diabetic Wound Healing.

Diabetic wounds are one of the debilitating complications that affect up to 20% of diabetic patients. Despite the advent of extensive therapies, the recovery rate is unsatisfactory, and approximately, 25% of patients undergo amputation, thereby demanding alternative therapeutic strategies. On the basis of the individual therapeutic roles of the miR-155 inhibitor and mesenchymal stem cells (MSC)-derived exosomes, we conjectured that the combination of the miR-155 inhibitor and MSC-derived exosomes would have synergy in diabetic wound healing. Herein, miR-155-inhibitor-loaded MSC-derived exosomes showed synergistic effects in keratinocyte migration, restoration of FGF-7 levels, and anti-inflammatory action, leading to accelerated wound healing mediated by negative regulation of miR-155, using an in vitro co-culture model and in vivo mouse model of the diabetic wound. Furthermore, treatment with miR-155-inhibitor-loaded MSC-derived exosomes led to enhanced collagen deposition, angiogenesis, and re-epithelialization in diabetic wounds. This study revealed the therapeutic potential of miR-155-inhibitor-loaded MSC-derived exosomes in diabetic wound healing and opened the doors for encapsulating miRNAs along with antibiotics within the MSC-derived exosomes toward improved management of chronic, nonhealing diabetic wounds.

Targeting specificity protein 1 with miR-128-3p overcomes TGF-ß1 mediated epithelial-mesenchymal transition in breast cancer: An in vitro study.

BACKGROUND: Specificity protein 1 (SP1) was found to play a critical role in the regulation of TGF-ß1 driven epithelial-mesenchymal transition (EMT). Recent clinical findings demonstrated a significant drop in the expression of miR-128-3p with the cancer progression in breast cancer patients. However, the impact of miR-128-3p on the SP1 expression in breast cancer remains unknown. Herein, we evaluated the role of miR-128-3p mimics in suppressing EMT of breast cancer cell lines by regulating the TGF-ß1/SP1 axis. METHODS: miR-128-3p interaction with SP1 was detected by in silico tools and dual-luciferase reporter assay. qPCR, western blot, and immunocytochemistry experiments were conducted for determining the expression levels of miR-128-3p and EMT markers with and without the treatment of miR-128-3p mimics. Further, to understand the effect of miR-128-3p mimics on cancer progression, experiments such as wound healing assay, transwell assay, adhesion assay, and cell cycle analysis were performed. RESULTS: A significant inverse relation between SP1 and miR-128-3p levels was found in MCF-7 and MDA-MB-231 cell lines. miR-128-3p overexpression impeded the SP1 mediated EMT markers in TGF-ß1 stimulated cells by inhibiting the SP1 nuclear function. Further, treatment with miR-128-3p mimics significantly reduced the migration, invasion and spreading capability of TGF-ß1 stimulated cells. Flow cytometry results showed the impeding role of miR-128-3p on the cell cycle progression. CONCLUSIONS: Upregulated miR-128-3p inhibited SP1, thereby limiting the TGF-ß1 induced EMT in MCF-7 and MDA-MB-231 cell lines for the first time. This study may pave the path to explore novel miRNA therapeutics for eradicating advanced breast cancer cases.

'Transfersome-embedded-gel' for dual-mechanistic delivery of anti-psoriatic drugs to dermal lymphocytes.

AIM: Develop a platform for co-delivering clobetasol propionate (CP) and cyclosporine (CyA) to the epidermis and dermis to treat psoriasis. METHODS: The transfersomes were prepared by thin-film hydration method. Transfersomes were characterised by dynamic light scattering and transmission electron microscope (TEM). Then, the gel stability, viscosity, pH, and spreadability were measured. Cytotoxicity of the CyA-loaded transfersome embedded in CP-dispersed gel (TEG-CyA-CP) was assessed on both human keratinocyte cell line (HaCaT) and Jurkat cells. In vitro cellular uptake and ex vivo dermal distribution was measured. The expression of inflammatory markers was assessed by reverse-transcription PCR (RT-PCR). RESULTS: Nanoscale (<150 nm) transferosomes with high CyA encapsulation efficiency (>86%) were made. TEG-CyA-CP demonstrated higher viscosity (4808.8 ± 12.01 mPas), which may help control dual drug release. Ex vivo results showed TEG-CyA-CP ability to deliver CyA in the dermis and CP in the epidermis. RT-PCR studies showed the optimised formulation helps reduce the tumour necrosis factor (TNF-α) and interleukin-1 (IL-1) levels to relieve psoriasis symptoms. CONCLUSION: The developed TEG-CyA-CP represents a promising fit-to-purpose delivery platform for the dual-site co-delivery of CyA and CP in treating psoriasis.

Tunneling Nanotube-Mediated Communication: A Mechanism of Intercellular Nucleic Acid Transfer.

Tunneling nanotubes (TNTs) are thin, F-actin-based membranous protrusions that connect distant cells and can provide e a novel mechanism for intercellular communication. By establishing cytoplasmic continuity between interconnected cells, TNTs enable the bidirectional transfer of nuclear and cytoplasmic cargo, including organelles, nucleic acids, drugs, and pathogenic molecules. TNT-mediated nucleic acid transfer provides a unique opportunity for donor cells to directly alter the genome, transcriptome, and metabolome of recipient cells. TNTs have been reported to transport DNA, mitochondrial DNA, mRNA, viral RNA, and non-coding RNAs, such as miRNA and siRNA. This mechanism of transfer is observed in physiological as well as pathological conditions, and has been implicated in the progression of disease. Herein, we provide a concise overview of TNTs' structure, mechanisms of biogenesis, and the functional effects of TNT-mediated intercellular transfer of nucleic acid cargo. Furthermore, we highlight the potential translational applications of TNT-mediated nucleic acid transfer in cancer, immunity, and neurological diseases.

Cyclo-RGD Truncated Polymeric Nanoconstruct with Dendrimeric Templates for Targeted HDAC4 Gene Silencing in a Diabetic Nephropathy Mouse Model.

Diabetic nephropathy (DN), a chronic progressive kidney disease, is a significant complication of diabetes mellitus. Dysregulation of the histone deacetylases (HDACs) gene has been implicated in the pathogenesis of DN. Hence, the HDAC-inhibitors have emerged as a critical class of therapeutic agents in DN; however, the currently available HDAC4-inhibitors are mostly nonselective in nature as well as inhibit multiple HDACs. RNA interference of HDAC4 (HDAC4 siRNA) has shown immense promise, but the clinical translation has been impeded due to lack of a targeted, specific, and in vivo applicable delivery modality. In the present investigation, we examined Cyclo(RGDfC) (cRGD) truncated polymeric nanoplex with dendrimeric templates for targeted HDAC4 Gene Silencing. The developed nanoplex exhibited enhanced encapsulation of siRNA and offered superior protection against serum RNase nucleases degradation. The nanoplex was tested on podocytes (in vitro), wherein it showed selective binding to the αvß3 integrin receptor, active cellular uptake, and significant in vitro gene silencing. The in vivo experiments showed remarkable suppression of the HDAC4 and inhibition in the progression of renal fibrosis in the Streptozotocin (STZ) induced DN C57BL/6 mice model. Histopathological and toxicological studies revealed nonsignificant abnormality/toxicity with the nanoplex. Conclusively, nanoplex was found as a promising tactic for targeted therapy of podocytes and could be extended for other kidney-related ailments.

MiR-155 Inhibitor-Laden Exosomes Reverse Resistance to Cisplatin in a 3D Tumor Spheroid and Xenograft Model of Oral Cancer.

Cisplatin resistance is one of the major concerns in the treatment of oral squamous cell carcinoma (OSCC). Accumulating evidence suggests microRNA (miRNA) dysregulation as one of the mediators of chemoresistance. Toward this, our previous study revealed the role of exosomal microRNA-155 (miR-155) in cisplatin resistance via downregulation of FOXO3a, a direct target of miR-155, and induction of epithelial-to-mesenchymal transition in OSCC. In the present study, we demonstrate the therapeutic potential of miR-155 inhibitor-laden exosomes in the sensitization of a cisplatin-resistant (cisRes) OSCC 3D tumor spheroid and xenograft mouse model. The cisRes OSSC 3D tumor spheroid model recapitulated the hallmarks of solid tumors such as enhanced hypoxia, reactive oxygen species, and secretory vascular endothelial growth factor. Further treatment with miR-155 inhibitor-loaded exosomes showed the upregulation of FOXO3a and induction of the mesenchymal-to-epithelial transition with improved sensitization to cisplatin in cisRes tumor spheroids and xenograft mouse model. Moreover, the exosomal miR-155 inhibitor suppressed the stem-cell-like property as well as drug efflux transporter protein expression in cisplatin-resistant tumors. Taken together, our findings, for the first time, established that the miR-155 inhibitor-loaded exosomes reverse chemoresistance in oral cancer, thereby providing an alternative therapeutic strategy for the management of refractory oral cancer patients.

MicroRNA-29b Modulates ß-Secretase Activity in SH-SY5Y Cell Line and Diabetic Mouse Brain.

Hyperglycemia is one of the major risk factors responsible for memory impairment in diabetes which may lead to Alzheimer's disease (AD) at a later stage. MicroRNAs are a class of non-coding RNAs that are found to play a role in diabetes. Downregulation of microRNA-29b in diabetes is well reported. Moreover, microRNA-29b is also reported to target the 3' UTR of ß-secretase (BACE-1) enzyme which is involved in the formation of amyloid-beta (Aß) in AD via cleavage of amyloid precursor protein (APP). Therefore, the present study was designed to elucidate whether microRNA-29b could be a link between diabetes and dementia. In the in vitro and in vivo diabetic model, we found downregulation of microRNA-29b due to hyperglycemia. After human microRNA-29b treatment, there was a significant improvement in the short-term and spatial memory in diabetic mice. Also, the human microRNA-29b treatment decreased oxidative stress and BACE-1 activity in diabetes. The present findings revealed that the downregulation of microRNA-29b in diabetes could be associated with memory impairment and increased BACE-1 activity. These results would give a future direction to study the role played by microRNAs in diabetes-associated memory impairment and hence aid in the development of therapeutics to treat the same.

Cognitive dysfunction: A growing link between diabetes and Alzheimer's disease.

Diabetes mellitus (DM) is a gradually rising metabolic disease which is currently affecting millions of people worldwide. Diabetes is associated with various complications like nephropathy, neuropathy, retinopathy, diabetic foot, cognitive impairment, and many more. Evidence suggests that cognitive dysfunction is a rising complication of diabetes which adversely affects the brain of patients suffering from diabetes. Age-related memory impairment is a complication having its major effect on people suffering from diabetes and Alzheimer's. Patients suffering from diabetes are at two times higher risk of developing cognitive dysfunction as compared with normal individuals. Multiple factors which are involved in diabetes related complications are found to play a role in the development of neurodegeneration in Alzheimer's. The problem of insulin deficiency and insulin resistance is well reported in diabetes but there are many studies which suggest dysregulation of insulin levels as a reason behind the development of Alzheimer's. As the link between diabetes and Alzheimer disease (AD) is deepening, there is a need to understand the plausible tie-ins between the two. Emerging role of major factors like insulin imbalance, advanced glycation end products and micro-RNA's involved in diabetes and Alzheimer's have been discussed here. This review helps in understanding the plausible mechanism underlying the pathophysiology of amyloid beta (Aß) plaque formation and tau hyperphosphorylation as well provides information about studies carried out in this area of research. The final thought is to enhance the scientific knowledge on this correlation and develop future therapeutics to treat the same.

Role of Omega-3 Fatty Acids and Butter Oil in Targeting Delivery of Donepezil Hydrochloride Microemulsion to Brain via the Intranasal Route: a Comparative Study.

In order to investigate the possible role of butter oil (BO) and omega-3 fatty acids-rich fish oil (O3FO) in the delivery of donepezil hydrochloride microemulsion (DH-ME) to the brain via intranasal route, the present study was conducted. DH:BO and DH:O3FO binary mixtures (9:1 to 1:9) were prepared by simple physical mixing and subjected to in vitro diffusion study. Ratios of DH:BO and DH:O3FO, which showed the highest diffusion, were selected for further development of microemulsion (ME). Globule sizes of DH-BO-ME and DH-O3FO-ME were found to be 87.66 ± 5.23 nm and 88.59 ± 8.23 nm, respectively. Nasal histopathological study and in vitro cytotoxicity study revealed the safety of the formulation. Higher percentage of nasal diffusion was found with DH-BO-ME (71.22 ± 1.21%) and DH-O3FO-ME (62.16 ± 1.23%) in comparison to DH-ME (59.69 ± 1.74%) and DH solution (55.01 ± 1.19%), which was further supported by in vitro cell permeability study. After intranasal administration, %bioavailability of drug in the rat brain (Sprague-Dawley rats)(on the basis of DH-ME IV) was higher with DH-BO-ME (313.59 ± 12.98%) and DH-O3FO-ME (361.73 ± 15.15%) in comparison to DH-ME (168.62 ± 6.60%) and DH solution (8.960 ± 0.23%). The results of ex vivo diffusion study and in vivo pharmacokinetic study suggested that BO and O3FO helped in enhancing the nasal permeability and the brain uptake of drug when administered intranasally.

Therapeutic restoration of miR-126-3p as a multi-targeted strategy to modulate the liver tumor microenvironment.

BACKGROUND: Impaired natural killer (NK) cell-mediated antitumor responses contribute to the growth of liver tumors. Expression of a disintegrin and metalloprotease 9 (ADAM9) increases shedding of membrane-bound major histocompatibility complex class I chain-related protein A and results in evasion from NK cell-mediated cytolysis. ADAM9 is also involved in angiogenesis and tumor progression and is a target of miR-126-3p, a tumor suppressor that is downregulated and alters tumor cell behavior in the liver and other cancers. We evaluated the restoration of miR-126-3p and modulation of the miR-126-3p/ADAM9 axis as a therapeutic approach to simultaneously enhance NK cell-mediated cytolysis while targeting both tumor cells and their microenvironment. METHODS: Precursor miRNAs were loaded into milk-derived nanovesicles to generate therapeutic vesicles (therapeutic milk-derived nanovesicles) for the restoration of functional miR-126-3p in recipient cancer cells. RESULTS: Administration of therapeutic milk-derived nanovesicles increased miR-126-3p expression and reduced ADAM9 expression in target cells and was associated with an increase in membrane-bound major histocompatibility complex class I chain-related protein A. This enhanced NK cell cytolysis in adherent tumor cells and in multicellular tumor spheroids while also impairing angiogenesis and modulating macrophage chemotaxis. Moreover, IV administration of therapeutic milk-derived nanovesicles with adoptive transfer of NK cells reduced tumor burden in orthotopic hepatocellular cancer xenografts in mice. CONCLUSION: A directed RNA therapeutic approach can mitigate NK cell immune evasion, reduce angiogenesis, and alter the tumor cell phenotype through the restoration of miR-126-3p in liver tumor cells. The pleiotropic effects elicited by this multi-targeted approach to modulate the local tumor microenvironment support its use for the treatment of liver cancer.

MiR-29b modulates DNA methylation in promoter region of miR-130b in mouse model of Diabetic nephropathy.

Epigenetic modifications play a role in Diabetic Nephropathy (DN). Downregulation of miR-29b leads to modulation of DNA methylation via DNA methyl transferases (DNMTs) and hence exaggerated renal fibrosis in DN. Therefore, the main aim of the study was to evaluate effect of miR-29b expression in vivo on DNMTs, renal fibrosis, glomerular and tubular damage as well as renal morphology in DN. In order to explore the role of miR-29b in DNA methylation of other miRNAs, methylation profiling study was performed. It revealed that miR-29b was involved in methylation on of miR-130b on the cytosine guanine dinucleotides rich DNA (CpG) island 1 located on promoter region. In conclusion, miR-29b expression was found to modulate DNA methylation via DNMTs and regulate methylation of miR-130b. The result of this study provides a future direction to unveil role of miRNA expression in DNA methylation and its consequent effect on other miRNAs in DN. Supplementary Information: The online version contains supplementary material available at 10.1007/s40200-023-01208-2.

Extracellular vesicle RNA signaling in the liver tumor microenvironment.

The tumor microenvironment (TME) in liver cancers such as hepatocellular cancer (HCC) consists of a complex milieu of liver tissue-resident cells, infiltrated immune cells, and secreted factors that collectively serve to promote tumor growth and progression. Intercellular crosstalk contributes to tissue homeostasis, and perturbations during injury, inflammation and tumorigenesis that are important for tumor progression. Extracellular vesicle (EV)-mediated transfer of a payload of RNA molecules that serve as an intercellular signaling is an important contributor to tissue homeostasis within the TME. Several types of RNA have been implicated in EV-mediated signaling. Biological processes that can be modulated by EV RNA signaling within the liver include tumor growth, invasion, metastasis, angiogenesis, and modulation of the immune cell activities. This mini-review describes the liver TME, and the biological effects of EV RNA-mediated signaling within the liver to highlight the role of EV RNA in intercellular communication.

Extracellular vesicle­mediated miR­126­3p transfer contributes to inter­cellular communication in the liver tumor microenvironment.

Extracellular vesicles (EVs) and their contents are gaining recognition as important mediators of intercellular communication through the transfer of bioactive molecules, such as non­coding RNA. The present study comprehensively assessed the microRNA (miRNA/miR) content within EVs released from HepG2 liver cancer (LC) cells and LX2 hepatic stellate cells (HSCs) and determined the contribution of EV miRNA to intercellular communication. Using both transwell and spheroid co­cultures of LC cells and HSCs, miR­126­3p within EV was established as a mediator of HSC to LC cell communication that influenced tumor cell migration and invasion, as well as the growth of multicellular LC/HSC spheroids. Manipulation of miR­126­3p either by enforced expression using pre­miR­126­3p or by inhibition using antimiR­126­3p did not alter tumor cell viability, proliferation or sensitivity to either sorafenib or regorafenib. By contrast, enforced expression of miR­126­3p decreased tumor­cell migration. Knockdown of miR­126­3p in tumor cells increased disintegrin and metalloproteinase domain­containing protein 9 (ADAM9) expression and in HSCs increased collagen­1A1 accumulation with an increase in compactness of multicellular spheroids. Within LC/HSC spheroids, ADAM9 and vascular endothelial growth factor expression was increased by silencing of miR­126­3p but diminished with the restoration of miR­126­3p. These studies implicate miR­126­3p in functional effects on migration, invasion and spheroid growth of tumor cells in the presence of HSCs, and thereby demonstrate functional EV­RNA­based intercellular signaling between HSCs and LC cells that is directly relevant to tumor­cell behavior.

Engineering Cell-Derived Nanovesicles for Targeted Immunomodulation.

Extracellular vesicles (EVs) show promise for targeted drug delivery but face production challenges with low yields. Cell-derived nanovesicles (CDNVs) made by reconstituting cell membranes could serve as EV substitutes. In this study, CDNVs were generated from mesenchymal stem cells by extrusion. Their proteomic composition, in vitro and in vivo toxicity, and capacity for loading RNA or proteins were assessed. Compared with EVs, CDNVs were produced at higher yields, were comprised of a broader range of proteins, and showed no detrimental effects on cell proliferation, DNA damage, or nitric oxide production in vitro or on developmental toxicity in vivo. CDNVs could be efficiently loaded with RNA and engineered to modify surface proteins. The feasibility of generating immunomodulatory CDNVs was demonstrated by preparing CDNVs with enhanced surface expression of PD1, which could bind to PD-L1 expressing tumor cells, enhance NK and T cell degranulation, and increase immune-mediated tumor cell death. These findings demonstrate the adaptability and therapeutic promise of CDNVs as promising substitutes for natural EVs that can be engineered to enhance immunomodulation.

Lung-on-chip microdevices to foster pulmonary drug discovery.

Respiratory diseases account for unprecedented mortality owing to a lack of personalized or insufficient therapeutic interventions. Fostering pulmonary research into managing pulmonary threat requires a potential alternative approach that can mimick the in vivo complexities of the human body. The in vitro miniaturized bionic simulation of the lung holds great potential in the quest for a successful therapeutic intervention. This review discusses the emerging roles of lung-on-chip microfluidic simulator devices in fostering translational pulmonary drug discovery and personalized medicine. This review also explicates how the lung-on-chip model emulates the breathing patterns, elasticity, and vascularization of lungs in creating a 3D pulmonary microenvironment.

Role of miRNAs in Cancer Diagnostics and Therapy: A Recent Update.

The discovery of microRNAs (miRNAs) has been one of the revolutionary developments and has led to the advent of new diagnostic and therapeutic opportunities for the management of cancer. In this regard, miRNA dysregulation has been shown to play a critical role in various stages of tumorigenesis, including tumor invasion, metastasis as well as angiogenesis. Therefore, miRNA profiling can provide accurate fingerprints for the development of diagnostic and therapeutic platforms. This review discusses the recent discoveries of miRNA- based tools for early detection of cancer as well as disease monitoring in cancers that are common, like breast, lung, hepatic, colorectal, oral and brain cancer. Based on the involvement of miRNA in different cancers as oncogenic miRNA or tumor suppressor miRNA, the treatment with miRNA inhibitors or mimics is recommended. However, the stability and targeted delivery of miRNA remain the major limitations of miRNA delivery. In relation to this, several nanoparticle-based delivery systems have been reported which have effectively delivered the miRNA mimics or inhibitors and showed the potential for transforming these advanced delivery systems from bench to bedside in the treatment of cancer metastasis and chemoresistance. Based on this, we attempted to uncover recently reported advanced nanotherapeutic approaches to deliver the miRNAs in the management of different cancers.

Engineered nanoplex mediated targeted miRNA delivery to rescue dying podocytes in diabetic nephropathy.

MicroRNAs (miRNA) is vital for gene expression regulation and normal kidney function. Mainly, miRNA-30a is responsible for the homeostasis of podocytes. In the diabetic nephropathic condition, miRNA-30a is directly and primarily suppressed by hyperglycemic kidney induced Notch signaling pathway leads to podocyte damage and apoptosis. Thus, transferring the exogenous miRNA-30a to podocytes might improve albuminuria as well as podocytes injury. The deprived stability, poor targetability, and low specificity in vivo are critical limitations to attain this objective. This investigation reports the specific and efficient delivery of miRNA-30a mimic via cyclo(RGDfC)-gated polymeric-nanoplexes with dendrimer templates to alleviate podocyte conditions. The nanoplexes able to protect RNase enzyme and to exhibit greater cellular uptake viaαvß3 receptor selective binding in HG treated podocytes. The nanoplexes up-regulated the expression level of miRNA-30a and repress the elevated Notch-1 signaling in HG exposed podocytes. The critical results of in vivo experimentation attribute marked suppression of Notch-1 in streptozotocin (STZ) induced diabetic C57BL/6 mice and reduced glomerular expansion and fibrosis in the glomerular area. Developed nanoplexes represents an efficient platform for the targeted delivery of exogenous miRNA to podocytes. The approach developed herein could be extrapolated to other gene therapeutics and other kidney-related diseases.

Structural features regulated photoluminescence intensity and cell internalization of carbon and graphene quantum dots for bioimaging.

Carbon-based nanostructures with nanometer dimensions have been identified as potential photoluminescence probes for bioimaging due to their biocompatibility, tunable bandgap, and resistance to photobleaching. However, the influence of structural features of carbon quantum dots (CQDs) and graphene quantum dots (GQDs) in bioimaging has not been explored previously. In the present investigation, we elucidated the mechanism of higher PL in GQDs as compared to CQDs as a function of their structural features. TEM and AFM studies revealed that CQDs were spherical (size ~5 nm), while GQDs showed zigzag edges (size ~3 nm). Further, XRD and NMR studies confirmed that CQDs and GQDs show amorphous and crystalline structures with greater sp2 clusters, respectively. While both the QDs demonstrated multicolor fluorescence against variable excitations with similar lifetime, GQDs showed 7-fold higher QY than CQDs. Bioimaging studies in 2D cell culture, 3D tumoroids, and in vivo suggested a greater intensity of fluorescence in GQDs than CQDs. Additionally, rapid cell internalization was observed in GQDs owing to their positive surface potential by heterogeneous atomic (N and S) doping. Moreover, both CQDs and GQDs have demonstrated better time dependent stability for fluorescence properties. Taken together, the proposed mechanism elucidates the greater PL intensity in GQDs due to quantum confinement effect, crystallinity, and surface edge effects and is a better candidate for bioimaging amongst the carbon family.

miR-29b attenuates histone deacetylase-4 mediated podocyte dysfunction and renal fibrosis in diabetic nephropathy.

PURPOSE: As epigenetic modifications like chromatin histone modifications have been suggested to play a role in the pathophysiology of Diabetic Nephropathy (DN) and are also found to be regulated by microRNAs. Our main purpose was to explore the role of microRNA in histone modulations associated with DN. There is downregulation of miR-29b due to advanced glycation end products in diabetes. Histone Deacetylase-4 (HDAC4) is amongst the histone modulators which promotes podocytes' impairment and upregulates transforming growth factor-1 (TGF-ß1) leading to renal fibrosis. Moreover, macrophage infiltration causes podocytes' apoptosis and IL-6 mediated inflammation. As miR-29b is downregulated in diabetes and HDAC4, TGF-ß1 and IL-6 could be the possible therapeutic targets in DN, our study was focussed on unveiling the role of miR-29b in modulation of HDAC4 and hence, in podocyte dysfunction and renal fibrosis in DN. METHODS: In silico analysis and luciferase assay were done to study the interaction between miR-29b and HDAC4. In-vitro DN model was developed in podocytes and miR-29b mimics were transfected. Also, podocytes were co-cultured with macrophage and miR-29b mimics were transfected. At the end, in-vivo DN model was generated in C57BL/6 J male mice and the effect of miR-29b mimics was reconfirmed. RESULTS: It was found that miR-29b targets the 3' untranslated region of HDAC4. In both in-vitro and in-vivo DN model, downregulation of miR-29b and subsequent increase in HDAC4 expression was observed. The miR-29b mimics suppressed podocytes' inflammation mediated through macrophages and attenuated HDAC4 expression, glomerular damage and renal fibrosis. CONCLUSION: This study concludes that miR-29b regulates the expression of HDAC4 which plays a role in controlling renal fibrosis and podocytes' impairment in DN.