Diosgenin alleviates D-galactose-induced oxidative stress in rats' brain and liver targeting aging and apoptotic marker genes.

The theory of aging is primarily concerned with oxidative stress caused by an imbalance in reactive oxygen species generation and cellular antioxidants. To alleviate the oxidative stress, we investigated the protective effect of diosgenin (DSG) for D-galactose (D-gal) using 20 and 40 mg of DSG/kg/day/orally for 42 days. The findings showed that D-gal caused brain and liver oxidative injuries by upregulating aging and oxidative markers. To counteract the oxidative stress caused by D-gal, DSG upregulated glutathione peroxidase-1, superoxide dismutase-1, and glutathione S-transferase-α. DSG also diminished the expression of p53, p21, Bcl-2-associated X protein, caspase-3, and mammalian target of rapamycin in brain and liver, as well as the build-up of ß-galactosidase. DSG, in a dose-dependent manner, decreased the oxidative aging effects of D-gal in brain and liver tissues through targeting of aging and apoptotic marker genes. Finally, it should be noted that consuming DSG supplements is a suggesting natural preventative agent that may counteract aging and preserve health through improvement of body antioxidant status and control aging associated inflammation and cellular apoptosis.

Reprogramming activated hepatic stellate cells by siRNA-loaded nanocarriers reverses liver fibrosis in mice.

We report on a novel strategy for treating liver fibrosis through reprogramming activated Hepatic Stellate Cells (aHSCs) into quiescent Hepatic Stellate Cells (qHSCs) using siRNA-loaded lipid nanoparticles (LNPs). The in vivo screening of an array of molecularly-diverse ionizable lipids identified two candidates, CL15A6 and CL15H6, with a high siRNA delivery efficiency to aHSCs. Optimization of the composition and physico-chemical properties of the LNPs enabled the ligand-free, selective, and potent siRNA delivery to aHSCs post intravenous administration, with a median effective siRNA dose (ED50) as low as 0.08 mg/Kg. The biosafety of the LNPs was confirmed by escalating the dose to 50-fold higher than the ED50 or by chronic administration. The recruitment of the novel LNPs for the simultaneous knockdown of Hedgehog (Hh) and Transforming Growth Factor Beta 1 (TGFß1) signaling pathways using an siRNA cocktail enabled the reversal of liver fibrosis and the restoration of the normal liver function in mice. Analysis of the key transcription factors in aHSCs suggested that the reprogramming of aHSCs into qHSCs mediated the therapeutic outcomes. The scalable ligand-free platform developed in this study as well as the novel therapeutic strategy reported herein are promising for clinical translation.

Human umbilical cord blood-derived mesenchymal stem cells restored hematopoiesis by improving radiation induced bone marrow niche remodeling in rats.

BACKGROUND: Functional hematopoiesis is governed by the bone marrow (BM) niche, which is compromised by radiotherapy, leading to radiation induced BM failure. The aim of this study was to demonstrate the radiation induced pathological remodeling of the niche and the efficacy of human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) in restoring hematopoiesis via improvement of the niche. METHODS: Thirty male Wistar rats were equally assigned to three groups: control (CON), irradiated (IR), and IR+hUCB-MSCs. Biochemical, histopathological and immunohistochemical analyses were performed to detect collagen type III and IV, Aquaporin 1+ sinusoidal endothelial cells and immature hematopoietic cells, CD11c+ dendritic cells, Iba1+ macrophages, CD9+ megakaryocytes, Sca-1+, cKit+, CD133 and N-cadherin+ hematopoietic stem and progenitor cells, CD20+, Gr1+ mature hematopoietic cells, in addition to ki67+ proliferation, Bcl-2+ anti-apoptotic, caspase-3+ apoptotic, TNF-α+ inflammatory cells. Histoplanimetry data were statistically analyzed using the one-way analysis of variance followed by the post hoc Duncan's test. Moreover, Pearson's correlation was used to assess the correlation between various parameters. RESULTS: In comparison to the IR group, the IR+hUCB-MSCs group showed restored cell populations and extracellular collagen components of the BM niche with significant increase in hematopoietic stem, progenitor, mature and proliferating cells, and a considerable decrease in apoptotic and inflammatory cells. Furthermore, highly significant correlations between BM niche and blood biochemical, histopathological, and immunohistochemical parameters were observed. CONCLUSION: hUCB-MSCs restored functional hematopoiesis through amelioration of the BM niche components via reduction of oxidative stress, DNA damage, inflammation, and apoptosis with upregulation of cellular proliferation.

Rutin attenuates D-galactose-induced oxidative stress in rats' brain and liver: molecular docking and experimental approaches.

Oxidative stress results from the imbalance between reactive oxygen species (ROS) production and antioxidant defence and is primarily involved in aging. The current study investigated the antioxidant activity of rutin in aging in rats induced by D-galactose (D-gal) for 42 days. Rutin was orally used at doses of 50 and 100 mg kg-1 daily. Results showed that D-gal induced oxidative alterations in the brain and liver recognized via upregulation of aging and oxidative markers. In contrast, rutin ameliorated the oxidative stress induced by D-gal by enhancing antioxidant markers such as superoxide dismutase-1, glutathione peroxidase-1, and glutathione S-transferase-α. Also, rutin significantly decreased the accumulation of ß-galactosidase and reduced the expression of p53, p21, Bcl-2-associated X protein (Bax), caspase-3 (CASP3), and mammalian target of rapamycin (mTOR) in brain and hepatic tissues. Rutin potentially attenuated these aging-related oxidative alterations in a dose-dependent manner. Moreover, rutin markedly reduced the increased immunohistochemical expression of ß-galactosidase, 8-hydroxy-2'-deoxyguanosine, calcium-binding adapter molecule 1, glial fibrillary acidic protein, Bax, and interleukin-6 and significantly increased Bcl2, synaptophysin, and Ki67. Furthermore, a molecular docking study revealed that rutin exhibited high affinity to rat and human caspases, PI3K/AKT/mTOR, and the IL-6 receptor. Finally, we can conclude that rutin supplementation can be a promising natural protective compound that could delay aging and maintain health.

Apitoxin alleviates methyl mercury-induced peripheral neurotoxicity in male rats by regulating dorsal root ganglia neuronal degeneration and oxidative stress.

Methylmercury (MeHg) toxicity is associated with extensive neuronal degeneration of dorsal root ganglia (DRG). This study aimed to assess the ameliorative effect of bee venom (BV) on methyl mercury chloride (MeHgCl)-induced peripheral neurotoxicity using DRGs in rats. Forty-eight adult male Sprague Dawley rats were allocated into four equal groups: G I: control (gavaged MilliQ water 1 ml/rat), G II: subcutaneously injected with BV (0.5 mg/kg b.wt), G III: gavaged MeHgCl (6.7 mg/kg b.wt), and G IV: received MeHgCl+BV. Dosing was done five times/week for 2 weeks. Ataxic behavior and visual impairments were significantly increased, whereas the movement behavior and motility gait were suppressed in the MeHgCl group. MeHgCl significantly decreased total antioxidant capacity (TAC) in DRG and significantly decreased the serum levels of glutathione (GSH), catalase (CAT), and superoxide dismutase (SOD). Tumor necrosis factor-alpha (TNF-α) and interleukin 1ß (IL-1ß) levels were significantly elevated, whereas interleukin 10 (IL-10) levels were significantly decreased in the MeHgCl group compared with the control group. DRGs of the MeHgCl-exposed rats showed pyknotic shrunken neurons with perineural vacuolations, demyelination of nerve axons, and proliferation of the satellite cells. MeHgCl significantly induced a higher positive index ratio of Iba-1, SOX10, neurofilament, pan-neuron, and vimentin immunostaining in the DRG. BV administration significantly mitigated the MeHgCl-induced alterations in oxidative stress-related indices. BV modified the immunostaining of Iba-1, SOX10, neurofilament, pan-neuron, and vimentin-positive index ratio in the DRG of the MeHgCl group. Our findings acknowledged that BV could enhance in vivo neuroprotective effects against MeHgCl-induced DRGs damage in male rats.

Self-homing nanocarriers for mRNA delivery to the activated hepatic stellate cells in liver fibrosis.

Herein, we report on the development of a platform for the selective delivery of mRNA to the hard-to-transfect Activated Hepatic Stellate Cells (aHSCs), the fundamental player in the progression of liver fibrosis. Using a microfluidic device (iLiNP), we prepared a series of lipid nanoparticles (LNPs) based on a diverse library of pH-sensitive lipids. After an in-depth in vivo optimization of the LNPs, their mRNA delivery efficiency, selectivity, potency, robustness, and biosafety were confirmed. Furthermore, some mechanistic aspects of their selective delivery to aHSCs were investigated. We identified a promising lipid candidate, CL15A6, that has a high affinity to aHSCs. Tweaking the composition and physico-chemical properties of the LNPs enabled the robust and ligand-free mRNA delivery to aHSCs in vivo post intravenous administration, with a high biosafety at mRNA doses of up to 2 mg/Kg, upon either acute or chronic administrations. The mechanistic investigation suggested that CL15A6 LNPs were taken up by aHSCs via Clathrin-mediated endocytosis through the Platelet-derived growth factor receptor beta (PDGFRß) and showed a pKa-dependent cellular uptake. The novel and scalable platform reported in this study is highly promising for clinical applications.

Engineered ε-decalactone lipomers bypass the liver to selectively in vivo deliver mRNA to the lungs without targeting ligands.

RNA drugs hold real potential for tackling devastating diseases that are currently resistant to small molecule drugs or monoclonal antibodies. However, since these drugs are unstable in vivo and unable to pass through cellular membranes their clinical realization is limited by their successful delivery to target sites. Herein we report on the design of a combinatorial library of polyester lipomers based on the renewable monomer, ε-decalactone (ε-DL), via organocatalytic ring-opening polymerization for mRNA delivery. The ε-DL lipomers showed a surprisingly efficient ability to target the lungs upon intravenous administration. Interestingly, most of the lipomers achieved functional EGFP expression in the lungs, while minimally transfecting hepatocytes and splenic cells. This simple approach for the design of biodegradable materials has the potential for the clinical translation of genetic medicines for the treatment of lung diseases.

Albendazole-loaded cubosomes interrupt the ERK1/2-HIF-1α-p300/CREB axis in mice intoxicated with diethylnitrosamine: A new paradigm in drug repurposing for the inhibition of hepatocellular carcinoma progression.

Hepatocellular carcinoma (HCC) is a leading cause of cancer related deaths worldwide. It was suggested that albendazole (ABZ) is a powerful inhibitor of several carcinoma types. However, the bioavailability of ABZ is very poor. Additionally, the mechanisms underlying the antitumor effects of ABZ may go beyond its tubulin-inhibiting activity. Therefore, we aimed to examine the effects of ABZ suspension (i.p. and p.o.) and ABZ-loaded cubosomes (LC) on the diethylnitrosamine-induced HCC in mice. ABZ-loaded nanoparticles exhibited a mean particle size of 48.17 ± 0.65 nm and entrapped 93.26 ± 2.48% of ABZ. The in vivo absorption study confirmed a two-fold improvement in the relative bioavailability compared with aqueous ABZ suspension. Furthermore, the oral administration of ABZ cubosomal dispersion demonstrated regression of tumor production rates that was comparable with ABZ (i.p.). ABZ relieved oxidative stress, improved liver function, and decreased necroinflammation score. The antiangiogenic activity was evident as ABZ effectively downregulated tissue expression of CD34, mRNA expression of CD309 and VEGF at the protein expression level. Besides, lower levels of MMP-9 and CXCR4 indicated antimetastatic activity. ABZ showed a considerable level of apoptotic activity as indicated by increased mRNA expression level of p53 and the increased Bax/BCL-2 ratio and active caspase-3. Additionally, Ki-67 expression levels were downregulated showing an antiproliferative potential. These protective effects contributed to increasing survival rate of diethylnitrosamine-treated mice. These effects found to be mediated via interrupting ERK1/2-HIF-1α-p300/CREB interactions. Therefore, our findings revealed that disrupting ERK1/2-HIF-1α-p300/CREB interplay might create a novel therapeutic target for the management of HCC.

Thymoquinone and Curcumin Defeat Aging-Associated Oxidative Alterations Induced by D-Galactose in Rats' Brain and Heart.

D-galactose (D-gal) administration causes oxidative disorder and is widely utilized in aging animal models. Therefore, we subcutaneously injected D-gal at 200 mg/kg BW dose to assess the potential preventive effect of thymoquinone (TQ) and curcumin (Cur) against the oxidative alterations induced by D-gal. Other than the control, vehicle, and D-gal groups, the TQ and Cur treated groups were orally supplemented at 20 mg/kg BW of each alone or combined. TQ and Cur effectively suppressed the oxidative alterations induced by D-gal in brain and heart tissues. The TQ and Cur combination significantly decreased the elevated necrosis in the brain and heart by D-gal. It significantly reduced brain caspase 3, calbindin, and calcium-binding adapter molecule 1 (IBA1), heart caspase 3, and BCL2. Expression of mRNA of the brain and heart TP53, p21, Bax, and CASP-3 were significantly downregulated in the TQ and Cur combination group along with upregulation of BCL2 in comparison with the D-gal group. Data suggested that the TQ and Cur combination is a promising approach in aging prevention.

Bee venom Apis mellifera lamarckii rescues blood brain barrier damage and neurobehavioral changes induced by methyl mercury via regulating tight junction proteins expression in rat cerebellum.

The objective of the current study is to investigate the protective effect of Egyptian bee venom (BV) against methyl mercury chloride (MMC) induced blood-brain barrier (BBB) damage and neurobehavioral changes. Eighty male Sprague-Dawley rats were randomly grouped into 1st control (C), 2nd BV (0.5 mg/kg S/C for14 days), 3rd MMC (6.7 mg/kg orally/14 days), and 4th MMC + BV group. MMC exposure significantly altered rat cognitive behavior, auditory startle habituation, and swimming performance, increased the exploratory, grooming, and stereotypic behavior. MMC significantly impaired BBB integrity via induction of inflammation, oxidative stress, and down-regulation of tight junction proteins genes (TJPs) mRNA expression levels: Occludin (OCC), Claudins-5 (CLDN5), Zonula occludens-1 (ZO-1), while up-regulated the transforming growth factor-beta (TGF-ß) mRNA expression levels. MMC revealed a significantly higher percentage of IgG positive area ratio, a higher index ratio of Iba1, Sox10, and ss-DNA, while index ratio of CD31, neurofilament, and pan neuron showed a significant reduction. Administration of BV significantly regulates the MMC altered behavioral responses, TJPs relative mRNA expression, and the immune-expression markers for specific neural cell types. It could be concluded for the first time that BV retains a promising in vivo protection against MMC-induced BBB dysfunction and neurobehavioral toxicity.

Ameliorative Role of Cerium Oxide Nanoparticles Against Fipronil Impact on Brain Function, Oxidative Stress, and Apoptotic Cascades in Albino Rats.

Fipronil (FIP) is an N-phenylpyrazole insecticide that is used extensively in public health and agriculture against a wide range of pests. Exposure to FIP is linked to negative health outcomes in humans and animals including promoting neuronal cell injury, which results in apoptosis through the production of reactive oxygen species (ROS). Therefore, the purpose of the current study was to investigate the neuroprotective effects of cerium oxide nanoparticles (CeNPs) on neuronal dysfunction induced by FIP in albino rats. Male rats were randomly classified into four groups: control, FIP (5 mg/kg bwt), CeNPs (35 mg/kg bwt), and FIP + CeNPs (5 (FIP) + 35 (CeNPs) mg/kg bwt), which were treated orally once daily for 28 consecutive days. Brain antioxidant parameters, histopathology, and mRNA expression of genes related to brain function were evaluated. The results revealed oxidative damage to brain tissues in FIP-treated rats indicated by the elevated levels of malondialdehyde (MDA) and nitric oxide (NO) levels and reduced activities of antioxidant enzymes such as superoxide dismutase (SOD) and glutathione peroxidase (GPx). On the other hand, the FIP's group that was treated with CeNPs showed decrease in MDA and NO levels and increase in SOD and GPx enzymes activity. Besides, FIP-treated rats showed decreased butyrylcholinesterase (BuChE) activity in comparison to the FIP + CeNPs group. Moreover, FIP caused up-regulation of the expression of neuron-specific enolase (NSE), caspase-3, and glial fibrillary acidic protein (GFAP) but down-regulation of B-cell lymphoma-2 (BCL-2) expression. But the FIP + CeNPs group significantly down-regulated the GFAP, NSE, and caspase-3 and up-regulated the gene expression of BCL-2. Additionally, the FIP-treated group of rats had clear degenerative lesions in brain tissue that was reversed to nearly normal cerebral architecture by the FIP + CeNPs treatment. Immunohistochemical examination of brain tissues of rats-treated with FIP showed abundant ionized calcium-binding adaptor molecule 1 (Iba-1) microglia and caspase-3 and apoptotic cells with nearly negative calbindin and synaptophysin reaction, which were countered by FIP + CeNPs treatment that revealed a critical decrease in caspase-3, Iba-1 reaction with a strong calbindin positive reaction in most of the Purkinje cells and strong synaptophysin reaction in the cerebrum and cerebellum tissues. Based on reported results herein, CeNPs treatment might counteract the neurotoxic effect of FIP pesticide via an antioxidant-mediated mechanism.

Aging-Related Functional and Structural Changes in Renal Tissues: Lesson from a Camel Model.

Renal aging is a progressive, physiological, and anatomical change that naturally occurs in all animal species. To date, no information is available concerning the aging-related structural and functional changes in camel kidneys. A total of 25 healthy male camels (14 aged 4­6 years and 11 aged 18­22 years) were included in this study. After the camels were slaughtered, samples were collected from all the camels' kidneys and prepared for histopathological, immunohistochemical, and gene expression evaluations. The most striking observation was the significant decline in the immunohistochemical abundance of podocin and the significant upregulation of smoothening in the aging camels' kidneys. However, the nonsignificant changes have reported for nephrin, calbindin, autophagy 5 (ATG5), aquaporin 1, and toll-like receptor 9. Furthermore, the mRNA expressions of sirtuin 1, superoxide dismutase 1, superoxide dismutase 2, peroxisome proliferator-activated receptor alpha, B-cell lymphoma 2 (Bcl-2), and erythropoietin were significantly decreased in the aging camels' kidneys. While the significant upregulation of Bcl-2-associated X protein and the nonsignificant increase in ATG5 expression levels were reported in the aging camels' kidneys. The present findings provide better understanding of the complex events and initiating factors of aging, allowing for the development of a future therapeutic strategy to preserve adequate renal function throughout life.

Mesenchymal stem cells enhance AQP1 expression in the sublingual salivary gland of ovariectomized menopausal rat model.

BACKGROUND: Ovariectomized menopausal rat model was used to investigate the effects of menopause on the sublingual salivary gland (SSG) and the potential therapeutic effect of human umbilical cord blood mesenchymal stem cells (hUCB-MSCs). METHODS: Thirty rats were equally divided into three groups: sham-operated (SHAM), ovariectomized (OVX), and ovariectomized stem cells injected (OVX+ hUCB-MSCs). Expressions of α-SMA, AQP1, Sca-1, PCNA, ssDNA, and caspase-3 were determined. Homing of hUCB-MSCs was detected by fluorescence microscopy and examination of immunostained sections for human CD105 and CD34 was performed. Morphometric data were statistically analyzed using the Kruskal-Wallis test followed by Scheffé's method. Correlation of AQP1 with Sca-1-positive sublingual stem cells was also analyzed. RESULTS: In the SSGs of the OVX group, ballooned mucus acinar cells, atrophied serous cells, and a decreased number and height of duct lining cells were observed. The interstitial spaces were edematous, and the blood vessels were congested. The significant decrease in the positive area % of α-SMA and AQP1, the number of Sca-1-positive sublingual stem cells, and proliferating cells was associated with a significant increase in apoptotic cells. The OVX+hUCB-MSCs group showed significant structural improvement, manifested by the normal appearance of mucus and serous acini, as well as the number and height of striated duct cells. A significant increase in the positive area % of α-SMA and AQP1 and the number of proliferating and Sca-1-positive sublingual stem cells was observed. Interestingly, a significantly positive Pearson's correlation between the area % of AQP1 and the number of Sca-1-positive sublingual stem cells was also recorded. CONCLUSION: Our results indicated a positive effect of hUCB-MSCs therapy for SSG pathology in a post ovariectomy rat model as evidenced by an improvement in the histologic architecture, upregulation of the immunostained area % of α-SMA and AQP1, increase in the number of Sca-1-positive sublingual stem cells and proliferating cells, and downregulation of apoptotic cells.

Unique localization of disseminated pancreas in the oesophagus of catfish (clarias gariepinus) with reference to sexual dimorphism.

BACKGROUND: The fish pancreas has been reported to be composed of two portions: compact and disseminated. However, little has been elucidated in catfish. The present study describes a unique localization of the disseminated pancreas in African catfish. METHODS: The sections were obtained and used for either routine histological examination following staining with haematoxylin and eosin (H & E), periodic acid-Schiff's, or were subjected to immunohistochemical staining for detection of both insulin-producing ß cells and glucagon-producing alpha cells. RESULTS: Our investigation showed that the pancreas of catfish consisted of both compact and disseminated portions. The compact pancreas was embedded in the mesenteric adipose tissue between the spleen, stomach and liver. However, the disseminated one showed unique localization in the tunica adventitia of the middle portion of the oesophagus. The pancreas consisted of two portions, exocrine and endocrine. Furthermore, in both types of pancreas, the female showed a significantly higher ratio for the endocrine islet area/pancreatic tissue area than that of the male and also a significantly higher ratio for both insulin- and glucagon-positive area/islet area in the female pancreas (compact and disseminated) than that of the male. IN CONCLUSION: The present study provides evidence on a unique localization of the disseminated pancreas in the oesophagus of catfish. Furthermore, we revealed sex-related difference in the endocrine portion in both pancreatic tissues with more development in the female. The study suggests that sex hormones could be contributed to such sexual dimorphism. However, further investigation is required to compare the degree of development during the spawning and resting seasons.

The potential ameliorative impacts of cerium oxide nanoparticles against fipronil-induced hepatic steatosis.

Fipronil (FIP) is a phenylpyrazole insecticide that is commonly used in agricultural and veterinary fields for controlling a wide range of insects, but it is a strong environmentally toxic substance. Exposure to FIP has been reported to increase the hepatic fat accumulation through altered lipid metabolism, which ultimately can contribute to nonalcoholic fatty liver disease (NAFLD) development. The present study aimed to examine the function of cerium oxide nanoparticles (CeNPs) in protecting against hepatotoxicity and lipogenesis induced by FIP. Twenty-eight male albino rats were classified into four groups: FIP (5 mg/kg/day per os), CTR, CeNPs (35 mg/kg/day p.o.), and FIP + CeNPs (5 (FIP) + 35 (CeNPs) mg/kg/day p.o.) for 28 consecutive days. Serum lipid profiles, hepatic antioxidant parameters and pathology, and mRNA expression of adipocytokines were assessed. The results revealed that FIP increased cholesterol, height-density lipoprotein, triacylglyceride, low-density lipoprotein (LDL-c), and very-low-density lipoprotein (VLDL-c) concentrations. It also increased nitric oxide (NO) and malondialdehyde (MDA) hepatic levels and reduced glutathione peroxidase (GPx) and superoxide dismutase (SOD) enzyme activities. Additionally, FIP up-regulated the fatty acid-binding protein (FABP), acetyl Co-A carboxylase (ACC1), and peroxisome proliferator-activated receptor-alpha (PPAR-α). Immunohistochemically, a strong proliferation of cell nuclear antigen (PCNA), ionized calcium-binding adapter molecule 1 (Iba-1), cyclooxygenase-2 (COX-2) reactions in the endothelial cells of the hepatic sinusoids, and increased expression of caspase3 were observed following FIP intoxication. FIP also caused histological changes in hepatic tissue. The CeNPs counteracted the hepatotoxic effect of FIP exposure. So, this study recorded an ameliorative effect of CeNPs against FIP-induced hepatotoxicity.

Novel lipid combination for delivery of plasmid DNA to immune cells in the spleen.

This study reports on the development of a novel lipid combination that permits the efficient and highly selective delivery of plasmid DNA (pDNA) to immune cells in the spleen. Using DODAP, an ionizable lipid that was previously thought to be inefficient for gene delivery, we show for the first time, that this ignored lipid can be successfully used for efficient and targeted gene delivery in vivo, but only when combined with DOPE, a specific helper lipid. Using certain DODAP and DOPE ratios resulted in the formation of lipid nanoparticles (LNPs) with a ~ 1000-fold higher gene expression, and this expression was specific for the spleen, making it the most spleen-selective system for transfection using pDNA. The developed DODAP/DOPE-LNPs target immune cells in the spleen via receptors for complement C3 and this pathway is critical for efficient gene expression. We hypothesize that the high spleen transfection activity of DODAP/DOPE-LNPs is caused by the promotion of gene expression associated with B cell activation via complement receptors. LNPs encapsulating tumor-antigen encoding pDNA showed both prophylactic and therapeutic anti-tumor effects. The optimized LNPs resulted in the production of different cytokines and antigen-specific antibodies as well as exerting antigen-specific cytotoxic effects. This study revives the use of DODAP in gene delivery and highlights the importance of using appropriate lipid combinations for delivering genes to specific cells.

Ultra-small lipid nanoparticles encapsulating sorafenib and midkine-siRNA selectively-eradicate sorafenib-resistant hepatocellular carcinoma in vivo.

Hepatocellular carcinoma (HCC) is a fatal disease with limited therapeutic choices. The stroma-rich tumor microenvironment hinders the in vivo delivery of most nanomedicines. Ultra-small lipid nanoparticles (usLNPs) were designed for the selective co-delivery of the cytotoxic drug, sorafenib (SOR), and siRNA against the Midkine gene (MK-siRNA) to HCC in mice. The usLNPs composed of a novel pH-sensitive lipid, a diversity of phospholipids and a highly-selective targeting peptide. A microfluidic device, iLiNP, was used and a variety of factors were controlled to tune particle size aiming at maximizing tumor penetration efficiency. Optimizing the composition and physico-chemical properties of the usLNPs resulted in an enhanced tumor accumulation, selectivity and in vivo gene silencing. The optimized usLNPs exerted potent gene silencing in the tumor (median effective dose, ED50~0.1 mg/Kg) with limited effect on the healthy liver. The novel combination synergistically-eradicated HCC in mice (~85%) at a surprisingly-low dose of SOR (2.5 mg/Kg) which could not be achieved via individual monotherapy. Toxicity studies revealed the biosafety of the usLNPs upon either acute or chronic treatment. Furthermore, the SOR-resistant HCC established in mice was eradicated by 70% using this approach. We conclude that our strategy is promising for potential clinical applications in HCC treatment.

Comparative Features of the Upper Alimentary Tract in the Domestic Fowl (Gallus gallus domesticus) and Kestrel (Falco tinnunculus): A Morphological, Histochemical, and Scanning Electron Microscopic Study.

The avian alimentary tract has evolved into different histologic structures to accommodate the physical and chemical features of several food types and flight requirements. We compared the esophagus, proventriculus, and gizzard of the domestic fowl, Gallus gallus domesticus (GGD) and kestrels, Falco tinnunculus (FT) using immunohistochemistry and scanning electron microscopy with various stains and lectins [Dolichos biflorus agglutinin (DBA) and Ricinus communis agglutinin I (RCA120)], and α-smooth muscle actin (α-SMA). The esophagus of GGD demonstrated thickened epithelium, muscularis mucosae, and inner circular longitudinal tunica muscularis layers; moderate outer longitudinal tunica muscularis layers; and a true crop. In contrast, the esophagus of FT showed a thin epithelium, no muscularis mucosae, moderate inner longitudinal and thick outer circular tunica muscularis layers, and no true crop. In the proventriculus, the nature of the secretion in GGD was neutral, but that of FT was acidic and neutral. In the gizzard, the muscle coat of GGD by α-SMA had no muscularis mucosae, unlike FT, which had muscularis mucosae. In summary, there are many histologic differences between GGD and FT to meet their different physiologic needs, such as feeding.

Pathological Alternations of Mediastinal Fat-Associated Lymphoid Cluster and Lung in a Streptozotocin-Induced Diabetic Mouse Model.

Diabetes is a devastating global health problem and is considered a predisposing factor for lung injury progression. Furthermore, previous reports of the authors revealed the role of mediastinal fat-associated lymphoid clusters (MFALCs) in advancing respiratory diseases. However, no reports concerning the role of MFALCs on the development of lung injury in diabetes have been published. Therefore, this study aimed to examine the correlations between diabetes and the development of MFALCs and the progression of lung injury in a streptozotocin-induced diabetic mouse model. Furthermore, immunohistochemical analysis for immune cells (CD3+ T-lymphocytes, B220+ B-lymphocytes, Iba1+ macrophages, and Gr1+ granulocytes), vessels markers (CD31+ endothelial cells and LYVE-1+ lymphatic vessels "LVs"), and inflammatory markers (TNF-α and IL-5) was performed. In comparison to the control group, the diabetic group showed lung injury development with a significant increase in MFALC size, immune cells, LVs, and inflammatory marker, and a considerable decrease of CD31+ endothelial cells in both lung and MFALCs was observed. Furthermore, the blood glucose level showed significant positive correlations with MFALCs size, lung injury, immune cells, inflammatory markers, and LYVE-1+ LVs in lungs and MFALCs. Thus, we suggest that the development of MFALCs and LVs could contribute to lung injury progression in diabetic conditions.

Hypoxia-Preconditioned Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells Mitigate Hypoglycemic Testicular Injury Induced by Insulin in Rats.

Hypoglycemia is a neglected metabolic disorder. Thus, we evaluated the protective effect of hypoxia-preconditioned human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) on hypoglycemic testicular injury. We examined 56 testes from 28 animals: 7 rats with insulin-induced hypoglycemia (HG group), 7 hypoglycemic rats which received an intratesticular injection of hUCB-MSCs (HG-MSC group), and 14 untreated control rats. Testosterone level, testicular catalase (CAT) activity, and malondialdehyde (MDA) level were analyzed. Immunostaining for specific testicular germ and somatic cell markers was performed. Proliferating and apoptotic cells were detected by anti-PCNA and anti-caspase-3, respectively. Morphometrical data were statistically analyzed. The hypoglycemic rats showed a significant decrease in testosterone level and CAT activity and a significant increase in MDA production. Examination of histological structure and protein expression of diverse germ cell markers revealed collapsed tubules that were lined by degenerated germ cells, decreased lactate dehydrogenase type C immune expression, as well as decreased proliferating and increased apoptotic cells number in hypoglycemic testes. Injection of MSCs improved testicular biochemical parameters, preserved germ cells and somatic cells, and decreased apoptosis. In conclusion, hypoxia-preconditioned hUCB-MSCs attenuate rat testicular injury caused by insulin-induced hypoglycemia. Avoidance and rapid management of hypoglycemia are necessary to avoid significant testicular injury.