Advanced probiotics: bioengineering and their therapeutic application.

The role of gut bacteria in human health has long been acknowledged and dysbiosis of the gut microbiota has been correlated with a variety of disorders. Synthetic biology has rapidly grown over the past few years offering a variety of biological applications such as harnessing the relationship between bacteria and human health. Lactic acid bacteria (LAB) are thought to be appropriate chassis organisms for genetic modification with potential biomedical applications. A thorough understanding of the molecular mechanisms behind their beneficial qualities is essential to assist the multifunctional medicinal sectors. Effective genome editing will aid in the creation of next-generation designer probiotics with enhanced resilience and specialized capabilities, furthering our knowledge of the molecular mechanisms behind the physiological impacts of probiotics and their interactions with the host and microbiota. The goal of this review is to provide a brief overview of the methods used to create modified probiotics with the scientific rationale behind gene editing technology, the mechanism of action of engineered probiotics along with their application to treat conditions like inflammatory bowel disease, cancer, bacterial infections, and various metabolic diseases. In addition, application concerns and future directions are also presented.

piRNAs in the human retina and retinal pigment epithelium reveal a potential role in intracellular trafficking and oxidative stress.

Long considered active only in the germline, the PIWI/piRNA pathway is now known to play a significant role in somatic cells, especially neurons. In this study, piRNAs were profiled in the human retina and retinal pigment epithelium (RPE). Furthermore, RNA immunoprecipitation with HIWI2 (PIWIL4) in ARPE19 cells yielded 261 piRNAs, and the expression of selective piRNAs in donor eyes was assessed by qRT-PCR. Intriguingly, computational analysis revealed complete and partial seed sequence similarity between piR-hsa-26131 and the sensory organ specific miR-183/96/182 cluster. Furthermore, the expression of retina-enriched piR-hsa-26131 was positively correlated with miR-182 in HIWI2-silenced Y79 cells. In addition, the lnc-ZNF169 sequence matched with two miRNAs of the let-7 family, and piRNAs, piR-hsa-11361 and piR-hsa-11360, which could modulate the regulatory network of retinal differentiation. Interestingly, we annotated four enriched motifs among the piRNAs and found that the piRNAs containing CACAATG and CTCATCAKYG motifs were snoRNA-derived piRNAs, which are significantly associated with developmental functions. However, piRNAs consisting of ACCACTANACCAC and AKCACGYTCSC motifs were mainly tRNA-derived fragments linked to stress response and sensory perception. Additionally, co-expression network analysis revealed cell cycle control, intracellular transport and stress response as the important biological functions regulated by piRNAs in the retina. Moreover, loss of piRNAs in HIWI2 knockdown ARPE19 confirmed altered expression of targets implicated in intracellular transport, circadian clock, and retinal degeneration. Moreover, piRNAs were dysregulated under oxidative stress conditions, indicating their potential role in retinal pathology. Therefore, we postulate that piRNAs, miRNAs, and lncRNAs might have a functional interplay during retinal development and functions to regulate retinal homeostasis.

A pilot-scale sustainable biorefinery, integrating mushroom cultivation and in-situ pretreatment-cum-saccharification for ethanol production.

Biomass pretreatment incurs 40% of the overall cost of biorefinery operations. The usage of mushroom cultivation as a pretreatment/delignification technique, and bio-ethanol production from spent mushroom substrates, after subsequent pretreatment, saccharification and fermentation processes, have been reported earlier. However, the present pilot-scale, entirely-organic demonstration is one of the very first biorefinery models, which efficiently consolidates: biomass pretreatment; in-situ cellulase production and saccharification; mushroom cultivation, thereby improving the overall operational economy. During pretreatment, the oyster mushroom, Pluerotus florida VS-6, matures into distinct substrate mycelia and fruiting bodies. Consequential variations in the kinetics of growth, biomass degradation/substrate utilization, oxygen uptake and transfer rates, and enzyme production, have been analyzed. Signifying the first-time usage of a biomass mixture, comprising vegetative waste and e-commerce packaging waste, the 30 day-long, bio-economical, non-inhibitor-generating, catabolite repression-limited, solid-state in-situ pretreatment-cum-saccharification, resulted in: 78% lignin degradation; 13.25% soluble-sugar release; 18.25% mushroom yield; 0.88 FPU/g.ds cellulase secretion. The in-situ saccharified biomass, when sequentially subjected to ex-situ enzymatic hydrolysis and fermentation, showed 37.35% saccharification, and a bio-ethanol yield of 0.425 g per g of glucose, respectively. Apart from yielding engine-ready bio-ethanol, the model doubles as an agripreneurial proposition, and encourages mushroom cultivation and consumption.

Therapeutic role of biogenic silver and gold nanoparticles against a DMH-induced colon cancer model.

Colorectal cancer (CRC) ranks third among fatal diseases afflicting mankind globally due to the shortage of primary detection methods and appropriate choice of drugs. Moreover, current treatments such as chemo drugs and radiotherapies create adverse effects and lead to drug resistance. In this context, recent advances in nanomedicine offer novel clinical solutions for colon cancer therapy. The current study denotes the therapeutic roles of biogenic Abutilon indicum silver and gold nanoparticles (AIAgNPs and AIAuNPs) against a 1, 2-dimethyl hydrazine (DMH)-induced CRC in Wistar rats. Following treatment of nanoparticles (NPs), the CRC rats showed great localization of AIAgNPs and AIAuNPs in colon tumors shown by ICP-OES, indicating their bioavailability. The AIAgNPs and AIAuNPs significantly enhanced cellular antioxidant enzyme levels including catalase, SOD, GSH, GPx and reduced lipid peroxidation (LPO) compared to the standard drug paclitaxel. AIAgNPs and AIAuNPs revealed significant protection against metastasis compared to paclitaxel shown in the histopathological study. The important CRC signaling molecules of the Wnt pathway, the ß-catenin and Tcf-4 levels were significantly downregulated in AIAgNPs and AIAuNPs treated CRC rats compared to paclitaxel. Furthermore, the expression levels of cleaved apoptotic caspase-9, -8, and - 3 and lamins were significantly upregulated in AIAgNPs and AIAuNPs treated CRC rats compared to paclitaxel. This preclinical study provides substantial insights into the anti-colon cancer roles of biogenic NPs and gives an idea for targeting different cancers.

Neuroprotective potential of Celastrus paniculatus seeds against common neurological ailments: a narrative review.

The most common human neurodegenerative diseases like Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD) etc. have been recognized to result from a complex interplay between genetic predisposition and defective cellular dynamics such as inappropriate accumulation of unfolded proteins, oxygen free radicals and mitochondrial dysfunction. The treatment strategies available today for these neurodegenerative ailments are only palliative and are incapable of restraining the progression of the disease. Hence, there is an immense requirement for identification of drug candidates with the ability to alleviate neuronal damage along with controlling progression of the disease. From time immemorial mankind has been relying on plants for treating varied types of dreadful diseases. Among the various medicinal plants used for treating various neurological ailments, Celastrus paniculatus (CP) popularly known as Jyotishmati or Malkangni is well known in the Ayurveda system of Indian Traditional Medicine whose seeds and seed oil have been used for centuries in treating epilepsy, dementia, facial paralysis, amnesia, anxiety, sciatica, cognitive dysfunctions etc. This review apart from specifying the phytochemical characteristics and traditional uses of C. paniculatus seeds and seed oil also exemplify the comprehensive data derived from various research reports on their therapeutic potential against some common neurological disorders.

Mechanism and Regulation of Autophagy in Cancer.

Autophagy, or self-eating, is a catabolic process that plays a crucial role in cellular homeostasis by carrying out bulk degradation of defective or superfluous proteins as well as worn-out organelles through a specialized structure, the autophagosome, which in turn fuses with the lysosome. Autophagy also alleviates cellular stress induced by nutrient deprivation, metabolic disturbance, hypoxia, and the like, by recycling intracellular constituents. This role of autophagy, to provide metabolic precursors especially upon starvation, might also contribute to the survival of cancer cells. The role of autophagy in cancer cells is ambiguous given that its downregulation or upregulation has been observed to depend on cancer stage and pathological grade. Autophagy has been found to exhibit a dual effect on tumorigenesis where it functions to suppress tumor progression by eliminating factors that cause genome instability while promoting survival of cancer cells under unfavorable conditions like therapeutic stress. This review aims to explain the mechanism, regulation, and the dual role of autophagy in cancer.

Green synthesized silver nanoparticles: Catalytic dye degradation, in vitro anticancer activity and in vivo toxicity in rats.

In this study, silver nanoparticles were synthesized (AgNPs) using aqueous rhizome extract of Acorus calamus (ACRE) and evaluated their in vitro anticancer activity and in vivo toxicity in a Wistar rat model. The synthesized AgNPs showed good catalytic activity against different organic pollutant dyes. In vitro cytotoxic effects of AgNPs were assessed in Hep2, COLO 205 and SH-SY5Y cells using MTT assay. Further, the apoptotic changes induced by AgNPs in more susceptible Hep2 cells were observed through AO/EB, DCFH-DA, Rhodamine 123, PI/DAPI staining, oxidative stress markers and Western blotting. In vivo toxicity study revealed substantial alterations in the levels of serum biochemical markers including AST, ALT, LDH and inflammatory markers such as TNF-α and IL-6 on day 29 when rats treated with AgNPs as compared to control, however, these levels were restored to normal at the end of washout period on day 89. No remarkable changes were observed in liver oxidative stress enzymes. ICP-OES analysis indicated bio-distribution of silver in spleen (5.67 µg/g) and liver (4.98 µg/g) in rats treated with 10 mg/kg b.w of AgNPs on day 29 and elimination of silver from all organs was observed at the end of washout period on day 89. Histopathological analysis revealed no significant changes in kidney, spleen, lungs, heart, testis and brain with 5 and 10 mg/kg b.w of AgNP. However, 10 mg/kg b.w of AgNPs showed moderate degree of cell swelling and vacuolar degeneration in liver and these alterations were reverted back to normal at the end of washout period. Findings from this study signify green synthesized AgNPs at low concentrations might be useful in many ways with ecofriendly nature.

In vivo bio-distribution, clearance and toxicity assessment of biogenic silver and gold nanoparticles synthesized from Abutilon indicum in Wistar rats.

This study reports the bio-distribution and clearance of Abutilon indicum silver and gold nanoparticles (AIAgNPs and AIAuNPs) in Wistar rats. Rats in different groups were orally administered with 5 and 10 mg/Kg BW of AIAgNPs and AIAuNPs (size 1-25 nm) for 28 days and few were maintained until 58 days of washout period. Serum biochemical parameters were not changed significantly at both doses of AIAuNPs and at lower concentration of AIAgNPs. But, with 10 mg/Kg BW of AIAgNPs rats showed elevated levels of AST, ALP and ALT on day 29, however, these levels were restored to normal after washout period. Liver oxidative stress markers were not altered with the treatment of AIAgNPs and AIAuNPs. ICP-OES analysis indicated bio-distribution of Ag and Au more in liver, kidney and spleen on day 29 and was found cleared on day 59. Histological analysis of nine vital organs indicated normal tissue architecture at both doses of AIAuNPs and lower dose of AIAgNPs. While the rats treated with higher dose of AIAgNPs showed mild liver sinusoid cell swelling on day 29, which also was recovered on day 59. Findings of this preclinical study indicate biocompatible nature of biogenic nanoparticles supporting their future biomedical applications.

Correction to: Licarin A induces cell death by activation of autophagy and apoptosis in non-small cell lung cancer cells.

The original version of this article unfortunately contained a mistake.

Licarin A induces cell death by activation of autophagy and apoptosis in non-small cell lung cancer cells.

Lung cancer has a relatively poor prognosis with a low survival rate and drugs that target other cell death mechanism like autophagy may help improving current therapeutic strategy. This study investigated the anti-proliferative effect of Licarin A (LCA) from Myristica fragrans in non-small cell lung cancer cell lines-A549, NCI-H23, NCI-H520 and NCI-H460. LCA inhibited proliferation of all the four cell lines in a dose and time dependent manner with minimum IC50 of 20.03 ± 3.12, 22.19 ± 1.37 µM in NCI-H23 and A549 cells respectively. Hence NCI-H23 and A549 cells were used to assess the ability LCA to induce autophagy and apoptosis. LCA treatment caused G1 arrest, increase in Beclin 1, LC3II levels and degradation of p62 indicating activation of autophagy in both NCI-H23 and A549 cells. In addition, LCA mediated apoptotic cell death was confirmed by MMP loss, increased ROS, cleaved PARP and decreased pro-caspase3. To understand the role of LCA induced autophagy and its association with apoptosis, cells were analysed following treatment with a late autophagy inhibitor-chloroquine and also after Beclin 1 siRNA transfection. Data indicated that inhibition of autophagy resulted in reduced anti-proliferative as well as pro-apoptotic ability of LCA. These findings confirmed that LCA brought about autophagy dependent apoptosis in non-small cell lung cancer cells and hence it may serve as a potential drug candidate for non-small cell lung cancer therapy.

Transition Metal Ion (Mn2+, Fe2+, Co2+, and Ni2+)-Doped Carbon Dots Synthesized via Microwave-Assisted Pyrolysis: A Potential Nanoprobe for Magneto-fluorescent Dual-Modality Bioimaging.

Heteroatom-doped carbon dots (C-dots) have captured widespread research interest owing to high fluorescence and biocompatibility for multimodal bioimaging applications. Here, we exemplify a rapid, facile synthesis of ethylenediamine (EDA)-functionalized transition metal ion (Mn2+, Fe2+, Co2+, and Ni2+)-doped C-dots via one-pot microwave (MW)-assisted pyrolysis at 800 W within 6 min using Citrus limon (lemon) extract as a carbon source. During MW pyrolysis, the precursor extract undergoes simultaneous carbonization and doping of metal ions onto C-dot surfaces in the presence of EDA. The EDA-functionalized transition metal ion-doped C-dots (i.e., Mn/C, Fe/C, Co/C, and Ni/C-dots) are collectively termed as TMCDs. The water-soluble TMCDs exhibited a size of 3.2 ± 0.485 nm and were enriched with amino and oxo functionalities and corresponding metal-oxide traces on the surfaces, as revealed from Fourier transfer infrared and X-ray photoelectron spectroscopy analyses. Interestingly, TMCDs demonstrated excitation-wavelength-dependent emission with brighter photoluminescence (PL) at 460 nm. Compared to pristine C-dots with a PL quantum yield (QY) of 48.31% and a fluorescence lifetime of 3.6 ns, the synthesized Mn/C, Fe/C, Co/C, and Ni/C-dots exhibited PL QY values of 35.71, 41.72, 75.07, and 50.84% as well as enhanced fluorescence lifetimes (τav) of 9.4, 8.6, 9.2, and 8.9 ns, respectively. The TMCDs significantly exhibited enhanced biocompatibility in human colon cancer cells (SW480) for fluorescence bioimaging and showed ferromagnetic and superparamagnetic behavior with vibrant T1-contrast ability. Interestingly, the maximum longitudinal (r1) relaxivity of 0.341 mM-1 s-1 was observed for Mn/C-dots in comparison to that of 3.1-3.5 mM-1 s-1 of clinically used Gd-DTPA magnetic resonance (MR)-contrast agent in vitro (1.5 T). Similarly, the maximum longitudinal relaxivity (r1) of 0.356 mM-1 s-1 was observed for Ni/C-dots (1.5 T) with respect to 4.16 ± 0.02 mM-1 s-1 attained for Gd-DTPA in vivo (8.45 T). Thus, the rapid, energy-efficient MW-assisted pyrolysis presents lemon extract derived, EDA-functionalized TMCDs with enhanced PL and efficient T1 contrast as potential magneto-fluorescent nanoprobes for dual-modality bioimaging applications.

Green synthesized nano silver: Synthesis, physicochemical profiling, antibacterial, anticancer activities and biological in vivo toxicity.

Green synthesis of nanoparticles using plants is gaining much interest in recent years. Anyway, intricate details on the role of phytochemicals involved in capping and stabilization of nanoparticles in diminishing toxicity and enhancing therapeutic potential are required. In this study, Ficus religiosa silver nanoparticles (FRAgNPs) were synthesized using Ficus religiosa leaf extract (FRLE) and characterized. The FRAgNPs showed good antibacterial activity and also cytotoxic effect in different cancer cell lines. Induction of apoptotic cell death was confirmed by various staining techniques, increased expression of cleaved caspases-8, 9, 3, lamin, PARP and oxidative stress markers in A549 and Hep2 cells. The in vivo studies performed in rats revealed significant increase in serum levels of AST, ALT, and LDH, TNF-α and IL-6 on day 29 following oral administration of FRAgNPs. However, these levels reverted back to normal at the end of wash out period on day 89. ICP-OES analysis revealed accumulation of silver in liver, brain and lungs on day 29 with respective concentration of 4.77, 3.94 and 3.043µg/g tissue. However, complete elimination of silver was observed on day 89. Histological analysis performed in vital organs indicated pathological changes only in liver which was also normalized after 89days.

Polyphenol stabilized colloidal gold nanoparticles from Abutilon indicum leaf extract induce apoptosis in HT-29 colon cancer cells.

Green synthesized gold nanoparticles have received substantial attention owing to their biomedical applications, particularly in cancer therapy. Although anticancer activities of green synthesized gold nanoparticles have been reported earlier, the underlying mechanism behind their anticancer activity is still to be understood. The present study, describes the green synthesis of Abutilon indicum gold nanoparticles (AIGNPs) from Abutilon indicum leaf extract (AILE) and their cytotoxic mechanism in colon cancer cells. Dimensions of spherical shaped AIGNPs were found to be in the range of 1-20nm as determined by TEM. GC-MS and FTIR analysis indicated the presence of polyphenolic groups in AILE, which might have been involved in the stabilization of AIGNPs. In vitro free radical scavenging analysis revealed the radical quenching activity of AIGNPs. Further, the AIGNPs exhibited cytotoxicity in HT-29 colon cancer cells with IC50 values of 210 and 180µg/mL after 24 and 48h. This was mediated through nuclear morphological changes and cell membrane damage as evidenced by acridine orange/ethidium bromide, propidium iodide and AnnexinV-Cy3 staining methods. Mechanism of the observed cytotoxicity of AIGNPs was explained on the basis of increased levels of reactive oxygen species and simultaneous reduction in cellular antioxidants, which might have caused mitochondrial membrane potential loss, DNA damage and G1/S phase cell cycle arrest. Expression of cleaved Caspase-9, Caspase-8, Caspase-3, Lamin A/C and PARP, provided the clues for the induction of intrinsic and extrinsic apoptosis pathways in AIGNPs treated HT-29 cells. The study provides a preliminary guidance towards the development of colon cancer therapy using green synthesized gold nanoparticles.

Neuroprotective effect of Valeriana wallichii rhizome extract against the neurotoxin MPTP in C57BL/6 mice.

Oxidative stress and inflammation are some of the contributing factors for dopaminergic neurodegeneration in Parkinson's disease (PD). Though Valeriana wallichii D.C. is known for its nervine activities its effect against PD is yet to be studied. This is the first report on the antioxidant and anti-inflammatory effect of V. wallichii rhizome extract (VWE) in MPTP induced PD mice. GC-MS analysis of VWE indicated the presence of phytoconstituents like isovaleric acid and acacetin. PD induced mice were treated orally with three different doses (50, 100 and 200mg/kg body weight (BW)) of VWE for 14 days and their behavioural changes were studied on days 0, 8, 13 and 21. The levels of striatal dopamine, mid brain tyrosine hydroxylase positive (TH(+)) cell count, TH protein expression, reactive oxygen species (ROS), lipid peroxidation (LPO), antioxidants and inflammatory cytokines were analysed. Mid brain glial fibrillary acidic protein (GFAP) expression was assessed by immunohistochemistry and western blotting. Also mid brain histopathological analysis was performed. VWE treatment significantly recuperated the altered behavioural test scores, striatal dopamine levels, mid brain TH(+) cell count and TH protein levels, increased GFAP expression and the histopathological changes observed in PD mice. Similarly, diminished levels of antioxidants, elevated levels of ROS, LPO and inflammatory cytokines were also significantly ameliorated following VWE treatment. The effective dose of VWE was found to be 200mg/kg BW. Conclusively, V. wallichii rhizome extract has the potential to mitigate oxidative stress and inflammatory damage in PD.

Biogenic silver nanoparticles from Abutilon indicum: their antioxidant, antibacterial and cytotoxic effects in vitro.

Green synthesis of silver nanoparticles using biological entities is gaining interest because of their potential applications in nano-medicine. Herein, we report the biological synthesis of Abutilon indicum silver nanoparticles (AIAgNPs) using aqueous Abutilon indicum leaf extract (AILE) and evaluation of their biological applications. TEM analysis revealed that the spherical biogenic AIAgNPs were found to be between 5 and 25 nm in size. The bioactive phyto-constituents such are condensed tannins of AILE were found to play a key role in the reduction and capping of AIAgNPs. The biological properties of AIAgNPs were premeditated as free radical scavenging activity, antibacterial effect and anti-proliferative activity. AIAgNPs were found to exhibit good free radical scavenging activities and the intense zone of inhibition displayed by them in six different pathogenic species indicate the potential antibacterial effect. Further, AIAgNPs showed a dose dependant anti-proliferative effect against COLO 205 (human colon cancer) and MDCK (normal) cells with an IC50 of 3 and 4 µg/mL and 100 and 75 µg/mL, respectively after 24 and 48 h. The morphological changes, chromatin condensation and membrane potential loss induced by AIAgNPs were evidenced by AO/EB and AnnexinV-Cy3 staining. The mitochondrial membrane potential (MMP) loss and G1/S transition cell cycle arrest in COLO 205 cells was evidenced in rhodamine123 staining and FACS analysis. The high levels of ROS as shown in DCF-DA staining could have played a major role in DNA fragmentation and eventually lead to apoptosis. The mode of action through the induction apoptosis by AIAgNPs in COLO 205 cells is exciting with promising application of nano-materials in biomedical research.

Biological activities of green silver nanoparticles synthesized with Acorous calamus rhizome extract.

Nanomedicine utilize biocompatible nanomaterials for diagnostic and therapeutic purposes. This study reports the synthesis of silver nanoparticles using aqueous rhizome extract of Acorus calamus (ACRE) and evaluation of antioxidant, antibacterial as well as anticancer effects of synthesized A. calamus silver nanoparticles (ACAgNPs). The formation of ACAgNPs was confirmed by UV-visible spectroscopy and their average size was found to be 31.83 nm by DLS particle size analyzer. Scanning electron micrograph (SEM) revealed spherical shape of ACAgNPs and energy dispersive spectroscopy (EDX) data showed the presence of metallic silver. Fourier transform infrared spectroscopy (FTIR) analysis indicated the presence of phenol/alcohol, aromatic amine and carbonyl groups in ACRE that were involved in reduction and capping of nanoparticles. ACRE and ACAgNPs exhibited substantial free radical quenching ability in various in vitro antioxidant assays performed in this study. ACAgNPs also displayed appreciable antibacterial activity against three different pathogenic bacteria and the growth kinetic study with Escherichia coli designated the inhibition of bacterial growth at the log phase. The cytotoxic effect of ACAgNPs was assessed by MTT assay in HeLa and A549 cells. The IC50 value of ACAgNPs respectively after 24 and 48 h was found to be 92.48 and 69.44 µg/ml in HeLa cells and in A549 cells it was 53.2 and 32.1 µg/ml. Apoptotic cell death in ACAgNPs treated cells was indicated by acridine orange/ethidium bromide (AO/EB) and annexinV-Cy3 staining techniques. Staining with propidium iodide (PI) and 4', 6-diamidino-2-phenylindole, dihydrochloride (DAPI) also confirmed nuclear changes such as condensation and fragmentation. Further, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay showed distribution of ACAgNPs treated cells in the late apoptotic stage. These findings emphasize that such biocompatible green nanoparticles with multifaceted biological activities may find their applications in the field of nanomedicine.