in vivo

Hydroxyapatite (HA), the main mineral component of bone, can be synthesized and utilized in the bone lesion treatments because of its high bioactivity and osteoconductive property. HA extraction from fish bones has received special attention given its low cost and easier extraction protocol compared to other sources. The present study compared the biocompatibility and bone repair of commercial nano hydroxyapatite (nHA) powder with that extracted from carp and human bones in vitro and in vivo.

First, nHA powders were prepared, and their physical and structural properties were studied using XRD, FTIR, FE-SEM and EDS analyses. Next, the powders were used to make porous scaffolds for which the physicochemical, structural, mechanical and biological properties were evaluated. The in vitro biocompatibility and osteogenic differentiation were tested on MC3T3-E1 cells, respectively, by MTT assay in three time periods and Alizarin red staining. Furthermore, the scaffolds were implanted after creation of critical-size lesions in the skulls of female rats, and the histological investigation was conducted by H&E staining at two time points.

The morphological and phase analyses showed the successful fabrication of porous nHA scaffolds with 60.68%, 61.38, and 63.27% for carp, human and commercial nHA scaffolds, respectively. The scaffolds showed different biodegradability behavior where the human nHA scaffolds degrade more rapidly. The results of mechanical tests indicated that the scaffolds made up of human extracted nHA powder had the lowest strength and stiffness (3.13 and 37.37 KPa, respectively). The strength and stiffness of the scaffolds fabricated by carp extracted and commercial nHA were 17.14 and 19.01 Kpa, and 251.79 and 140.49 Kpa, respectively. The MTT test results showed that the greatest cell viability rate was in the carp nHA scaffolds after 10 days (146.08%). Moreover, the AR staining indicated the formation of mineralized nodules caused by the scaffolds in all groups. However, the mineralization seemed to be superior in human, and carp extracted groups. Furthermore, in vivo in all three groups bone repair occurred at the critical-size lesion sites, while scaffolds biodegradation was also observed. The scaffolds made up of carp and human nHA exhibited the highest rate of ossification and maturation of bone tissue among different scaffolds after 8 weeks. The rate of tissue response to these scaffolds was higher than the scaffolds made of commercial nHA after 4 and 8 weeks, postoperatively.

The carp extracted nHA scaffolds perform comparable to human extracted nHA, and may be used for clinical applications.

Graphene and its derived forms have surfaced as promising substances for a wide range of technological and biomedical purposes. However, it is crucial to evaluate their safety and potential risks to ensure their safe use. This systematic review examines the present state of knowledge regarding the toxicity of graphene oxide (GO) through in-vivo, in-vitro, and other species studies. The aim of this present research was to study toxicity outcomes of GO-coated materials. The literature search was conducted and most important electronic databases (20 studies) were checked and selected for the present study. The findings underscore the need for cautious consideration of GO’s potential risks, especially at high concentrations and prolonged exposures. Continued research efforts are essential to gain a deeper understanding of the underlying mechanisms and to develop appropriate safety guidelines for the utilization of GO in various applications.

 Warfarin is one of the most widely used anticoagulants that functions by inhibiting vitamin K epoxide reductase. Warfarin overdose, whether intentional or unintentional, can cause life-threatening bleeding. Here, we present a novel warfarin adsorbent based on mesoporous silica that could serve as an antidote to warfarin toxicity.

 Amino-functionalized mesoporous silica (MS-NH2 ) was synthesized based on the co-condensation method through a soft template technique followed by template removal. The prepared structure and functional group were studied by Fourier transform infrared spectroscopy (FT-IR), and X-ray diffraction (XRD). Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) checked the morphology. The capacity of MS-NH2 in the adsorption of warfarin was evaluated in vitro, at pH=7.4 and pH=1.2. In vivo evaluations were performed in control and warfarin-overdosed animal models. Overdosed animals were treated with MS-NH2 by oral gavage. Biomarkers of organ injury were assessed in animal serum.

 The MS-NH2 was relatively uniform, spherical with defined diameters (400 nm) and porous structure. Synthesized particles had a large surface area (1015 m2 g-1) and mean pore diameter of 2.4 nm which led to considerable adsorption capacity for warfarin 1666 mg/g. In vivo studies revealed that oral administration of MS-NH2 in mice poisoned with warfarin caused a significant difference (P<0.05) in the International Normalized Ratio (INR) and prothrombin time (PT). Moreover, the warfarin with MS-NH2 group demonstrated a notable decrease in biomarkers associated with tissue damage, such as bilirubin, lactate dehydrogenase (LDH), alanine aminotransferase (ALT), and aspartate aminotransferase (AST).

 The results confirm that MS-NH2 administration can be an effective treatment for warfarin toxicity and could potentially mitigate the adverse effects of warfarin poisoning.

Toxoplasma gondii transmission can occur during pregnancy if the mother contracts the infection for the first time. Treatment strategies include the use of antimicrobial medications and providing supportive care. Spiramycin is commonly used to treat toxoplasmosis in pregnant women and to hinder the disease's transmission. However, its ability to treat the fetus is questionable due to its limited capacity to cross the placental barrier. Additionally, economic constraints and sanctions may impede access to this medication.

Consequently, in search of an effective treatment, for the first time in Iran, the effectiveness of clindamycin in preventing abortion and vertical transmission of the PRU strain of T. gondii infection in pregnant mice was evaluated.

On the twelfth day of gestation, pregnant mice were exposed to T. gondii and subsequently received treatment with either clindamycin or spiramycin. This resulted in the establishment of four distinct groups: A normal control group, an infected group without treatment, an infected group treated with clindamycin, and another infected group treated with spiramycin. Following these interventions, a series of parasitological evaluations (including microscopic examination and realtime PCR), histopathological evaluations, and immunological assessments were conducted.

The findings showed a significant reduction in the number of cysts in the eye and brain (ranging from 77.32% to 90.72%) among the groups treated with clindamycin and spiramycin compared to the control group. Furthermore, treatment with clindamycin, like treatment with spiramycin, was able to suppress inflammatory changes, prevent cell death, and reduce vascular cuffs in the brain, as well as decrease bleeding, placental thrombosis, and the accumulation of inflammatory cells in the placenta. Clindamycin was also effective in diminishing retinal folds, tiny retinal bleeds, and cell vacuolation in eye tissues. Immunologically, treatment in both the spiramycin and clindamycin groups resulted in a decrease in the level of the cytokine TNF-α, indicating an increase in the cellular immune response. In addition, increased levels of IL-10 in the treated infected groups could contribute to the reduction of TNF-α production.

Typically, spiramycin is the first choice for treating congenital toxoplasmosis, but clindamycin can be a useful substitute or additional treatment when resistance to primary medications occurs, when there is intolerance, or when access to the main drugs is restricted.

Cancer is the second leading cause of death worldwide, surpassed only by cardiovascular diseases. This study investigated the anticancer effects of recombinant Clostridium α-toxin on breast cancer, both in vitro and in vivo. The entire coding sequence of a codon-optimized α-toxin was designed, cloned into the pET28a (+) vector, and expressed as recombinant α-toxin in Escherichia coli (E. coli) BL 21(DE3) cells transformed with the recombinant plasmid. The recombinant α-toxin was purified using Ni²⁺ affinity chromatography, and its accuracy and purity were confirmed through sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis. The anticancer effects of purified α-toxin were then assessed in vitro and animal models against MCF-7 breast cancer cells. Protein analysis confirmed the presence of a 48 kDa band corresponding to the recombinant α-toxin. Additionally, the IC₅₀ values of α-toxin against MCF-7 cells at 24, 48, and 72 h were 407.3±2.392 μg/mL, 287.3±5.411 μg/mL, and 258.1±4.671 μg/mL, respectively. In vivo, results demonstrated a significant reduction in mean cancer nodule size following α-toxin treatment (p<0.001). These findings suggest that α-toxin may serve as a promising candidate for breast cancer therapy.

New cancer therapies based on Escherichia coli (E. coli) have gained significant interest recently. E. coli with genetically modified genes were investigated in this study for their potential to treat cancers.  

The present systematic review was conducted to gather relevant literature on E. coli-based cancer therapies. The current study searched several databases for preclinical studies and early-phase clinical trials. Those studies included in-vitro and in-vivo evaluations of genetically engineered E. coli used for cancer therapy. Furthermore, the current study evaluated the potential of E. coli-based therapy to treat cancer in combination with other therapies and using a personalized approach.  

After meticulously reviewing 13,064 publications, 301 studies were included for quantitative analysis, following a screening process, including 44 articles. Live tumor-targeting bacteria have the potential to revolutionize cancer therapeutics, according to the review. Despite the challenges associated with conventional cancer treatments, E. coli offers an alternative strategy that can accumulate and increase within tumors. A variety of anticancer agents can be carried by E. coli via genetic manipulation and synthetic bioengineering, making them ideal carriers for tailored therapeutic approaches. Researchers found they could be used as a monotherapy or combination therapy, presenting a multifaceted solution for enhanced clinical outcomes. Several clinical trials with E. coli targeting tumors are underway, demonstrating that theoretical promise has been translated into practical application.  

To conclude, live tumor-targeting bacteria may be able to address the limitations of existing cancer treatments. The selectivity of their antitumor immune responses, programmability, and ability to induce antitumor immune responses indicate a significant advancement. Despite these challenges, ongoing clinical trials suggest a tangible shift in how E. coli can be integrated into cancer treatment regimens. More research and development are needed to utilize these new targeted anticancer strategies to their full potential.

Toxoplasma gondii is one of the most common parasites worldwide. It is of great importance to identify new potential drugs that are effective and less harmful in pregnant women and newborns. We investigated nanoemulsion miltefosine (NEM) in treating experimental acute and chronic toxoplasmosis.

A combination of triacetin, Tween 80, and ethanol (1:2) was used for the development of NEM formulations. The size of NEM was calculated to be 17.463 nm by DLS and TEM. To investigate the performance of miltefosine (MLF), NEM, sulfadiazine (SDZ), and pyrimethamine (PYR) (positive control) in vivo, acute toxoplasmosis was induced in mice by an intraperitoneal injection of RH strain tachyzoites. After five days, the mice were examined for the number and condition of tachyzoites and histopathological changes in the liver and spleen. Chronic toxoplasmosis was investigated in rats and the number and size of brain cysts along with histopathological changes were assessed in different groups.

The results of the in vivo assessment of drugs in acute toxoplasmosis showed the following order regarding a decrease in the number of tachyzoites and an increase in survival rate: SDZ&PYR > NEM > MLF. The effects of drugs on chronic toxoplasmosis showed a significant effect of NEM (50%) on reducing the number of cysts compared to SDZ&PYR (10%) and MLF (12%) and reducing the size of NEM brain cysts (21%) compared to SDZ&PYR (5 %) and MLF (8%).

Increasing the penetration of NEM through the blood-brain barrier (BBB) and subsequently reducing the number and size of T. gondii tissue cysts is a promising new drug in treating chronic toxoplasmosis.

The hydatid cyst (cystic echinococcosis) is the larval stage of Echinococcus granulosus sensu lato, which accounts for significant zoonotic infections worldwide.

This study aimed to evaluate the in vitro and in vivo efficacy of the essential oil derived from Thymbra spicata L. (TSEO) against protoscoleces (PSCs) and hydatid cysts of E. granulosus .

The components of TSEO were characterized using gas chromatography-mass spectrometry (GC-MS). The in vitro effects of TSEO on PSCs were determined using the eosin exclusion test. The effect of TSEO on caspase-3 gene expression and exterior ultrastructure of PSCs was investigated using real-time PCR and scanning electron microscopy (SEM), respectively. In vivo effects of TSEO at doses of 20, 30, and 40 mg/kg/day were also studied in mice with hydatid cysts.

Gas chromatography-mass spectrometry analysis showed that the main compounds in TSEO were carvacrol (65.88%), γ-terpinene (9.71%), and p-cymene (7.82%), respectively. The results showed the highest lethality of TSEO at 30 µg/mL after 60 minutes of exposure. The IC50 value was 18.60 µg/mL after 60 minutes of exposure to TSEO. After exposing PSCs to TSEO, the expression level of the caspase-3 gene increased over time with increasing concentration (P < 0.05). Scanning electron microscopy images of the cyst treated with 50 µL/mL of TSEO showed ultrastructural damage, rostellar disorganization, alterations in the teguments, and deformation of the cyst structure. In the in vivo assay, it was found that the average number, size, and weight of hydatid cysts decreased significantly (P < 0.05) after treatment with TSEO. The maximum efficacy was observed after treatment with TSEO at a dose of 40 mg/kg, resulting in a significant decrease in the number, weight, and size of hydatid cysts by 9.6 ± 1.51, 2.64 ± 0.39 g, and 0.18 ± 0.022 mm, respectively.

The study findings confirmed the promising in vitro and in vivo effects of TSEO against hydatid cyst infection. Considering the possible mechanisms, TSEO provoked cell wall damage and induced apoptosis. However, more studies are needed to confirm these findings and clarify the precise mechanisms of action.

 Receptor-mediated transcytosis (RMT) is a more specific, highly efficient, and reliable approach to crossing the blood-brain-barrier (BBB) and releasing the therapeutic cargos into the brain parenchyma.

 Here, we introduced and characterized a human/mouse-specific novel leptin-derived peptide using in silico, in vitro and in vivo experiments.

 Based on the bioinformatics analysis and molecular dynamics (MD) simulation, a 14 amino acid peptide sequence (LDP 14) was introduced and its interaction with leptin-receptor (ObR) was analyzed in comparison with an well known leptin-derived peptide, Lep 30. MD simulation data revealed a significant stable interaction between ligand binding domains (LBD) of ObR with LDP 14. Analyses demonstrated suitable cellular uptake of LDP 14 alone and its derivatives (LDP 14-modified G4 PAMAM dendrimer and LDP 14-modified G4 PAMAM/pEGFP-N1 plasmid complexes) via ObR, energy and species dependent manner (preferred uptake by human/mouse cell lines compared to rat cell line). Importantly, our findings illustrated that the entry of LDP 14-modified dendrimers in hBCEC-D3 cells not only is not affected by protein corona (PC) formation, as the main reason for diminishing the cellular uptake, but also PC per se can enhance uptake rate. Finally, fluorescein labeled LDP 14-modified G4 PAMAM dendrimers efficiently accumulated in the mice brain with lower biodistribution in other organs, in our in vivo study.

 LDP 14 introduced as a novel and highly efficient ligand, which can be used for drugs/genes delivery to brain tissue in different central nervous system (CNS) disorders.

 This study evaluated the cytotoxicity of four bioceramic root canal sealers (RCSs) in vivo. The embryonic zebrafish characteristics, such as mortality, survival, hatching, and general morphology, served as the parameters for assessing cytotoxicity.

 The RCSs, namely GuttaFlow Bioseal, MTA Fillapex, CeraSeal Bioceramic, and iRoot SP, were mixed according to the manufacturer’s guidelines. The extract solution was prepared by immersing the set RCS into 1X dilution of E3 solution. Then, the extract solution was delivered into a Petri dish where zebrafish embryos were allowed to develop. Cytotoxicity was evaluated 24, 48, 72, and 96 hours after fertilization.

 The Kruskal-Wallis test showed that except for GuttaFlow Bioseal, the mortality, survival, and hatching of zebrafish embryos for the remaining three bioceramic RCSs were significantly different from the negative controls (P<0.05). Significant differences were also evident in the mortality, survival, and hatching of zebrafish embryos between GuttaFlow Bioseal and three other RCSs (P<0.05).

 GuttaFlow Bioseal was less cytotoxic than other bioceramics RCSs; MTA Fillapex, CeraSeal Bioceramic root canal sealer, and iRoot SP root canal sealer exhibited comparable cytotoxicity.

In vitro and in vivo researches have shown that silver nanoparticles have more antimicrobial properties with a lower concentration than antifungal agents against candida vaginitis. Therefore, this study evaluated the therapeutic effect of silver nanoparticles (Nivasha spray15ppm) compared to clotrimazole 1% vaginal cream on candida vaginitis.

In this clinical trial study, 110 women with confirmed candida vaginitis randomly were divided into test (n=58) and control (n=52) groups. Silver nanoparticles spray with an applicator (Nivasha 15 ppm), and clotrimazole 1% were administered to test and control groups, respectively. Then, within ten days, post-intervention checkup and patient self-reported for treatment results were recorded in checklists and the data were analyzed statistically.

The improvement rate in test group (98.0%) was 1.44 times higher than in control (67.9%). Moreover, disease symptoms after the intervention (including unusual secretions, itching and burning, redness) in test group were significantly less than in the control, but there was no significant difference in the ratio of edema in two groups )p=0.071(. Furthermore, the average recovery time (days) of all symptoms in test group was lower than control (p<0.05). Finally, the rate of patients' satisfaction with the treatment process in the test group (76.9%) was more than control (46.6%) (p=0.004).

Nivasha spray had more effectiveness compared to the clotrimazole 1%. Therefore, it can be used as an alternative drug in the treatment of Candida vaginitis.

Burn is one of the prominent causes of death around the world, however drug discovery attempts for burn healing has not been entirely successful. Aloe arborescens (A. arborescens), is effective in the burning wounds healing and growth inhibition of bacterial pathogens. Our objective was to assess the wound healing and antibacterial effects of A. arborescens in vivo.

Thirty healthy Wistar rat animals were enrolled. The treatment process continued for 21 days and sampling was conducted on days 14 and 21 and the tissue slides were sent to the pathology laboratory for testing. The bactericidal activity of A. arborescens extract was evaluated using the disc diffusion method.

A. arborescens demonstrated a significant effect on the healing of burn wounds. Furthermore, the antibacterial effects of the A. arborescens extract against Gram-negative (Escherichia coli, Pseudomonas aeruginosa) was significantly higher than that against Gram-positive (Staphylococcus aureus, Bacillus cereus) bacterial species.

In conclusion, this study indicated that A. arborescens extract had an improving effect on the healing process of third degree burns without toxicity to the tissue.

As the use of Gentamicin became more widespread, the drug’s harmful effects, particularly nephrotoxicity, became increasingly well-known. Antibacterial and anti-inflammatory properties have long been associated with Mirazid. The goal of this research was to find out more about frameworks for the protection of Mirazid against nephrotoxicity triggered by Gentamicin.

Three groups of albino male rats were created; the normal group received only saline. In the second group, nephrotoxicity was produced for 10 days with Gentamicin (100 mg/kg; i.p.). In the third group; Mirazid (10 mg/kg; p.o.) was administered for 10 days before receiving Gentamicin. This was done to investigate the kidney/body weight index, serum creatinine, urea, lactate dehydrogenase (LDH), malondialdehyde (MDA), and Glutathione (GSH) levels. Moreover, immunohistochemical staining was done to study Jun N- terminal kinase 1 (JNK1), inducible nitric oxide synthase (iNOS), and caspase3 expressions along with histopathological changes. Additionally, a molecular docking study was performed for the seventeen isolated and identified compounds from myrrh, JNK1 is inhibited by an oleo-gum resin derived from the Commiphora species of plants (Burseraceae).

The Gentamicin group showed an increase in kidney/ body weight index, serum creatinine, urea, LDH, and MDA, while decreasing GSH levels. Furthermore, immunohistochemical staining revealed increased JNK1, iNOS, and caspase3 expressions along with histopathological changes. All of these indicators were significantly reduced by mirazid, which also restored oxidant/antioxidant hemostasis. Furthermore, the histological architecture of tissues has been significantly conserved. Concerning the docking study, the isolated compound (12) was found to be superior to the co-crystallized inhibitor (18) with a binding score of -7.19 kcal/mol compared to -6.95, respectively.

Mirazid was found to be a potential method for suppressing the nephrotoxicity caused by Gentamycin by inhibiting the JNK1/ iNOS pathways, therefore preserving kidney function. The antioxidant, anti-inflammatory, and anti-apoptotic properties of mirazid are thought to be responsible for its preventive efficacy.

Toxoplasma gondii is a zoonotic parasite of increasing concern to humans and animals. Considering the side effects of drugs used to treat toxoplasmosis, it is essential to find alternative drugs.

In this study, colchicine and propranolol at four concentrations (1, 5, 10, and 15 µg/mL) were added to the RPMI medium containing peritoneal macrophages and incubated for 60 min, Then tachyzoites were added to the medium, and the efficacy rates of colchicine and propranolol in inhibiting tachyzoites entry into macrophages were evaluated after 30 and 60 min. For in vivo assay, one group received no drugs, and the second group was treated with colchicine and propranolol at different concentrations for different durations.

The in vitro experiment showed that treatment with 15 mg/mL of colchicine and propranolol for 60 min following tachyzoites addition was the most efficient method to inhibit tachyzoites penetration, indicating the efficacy rates of 80.20%±1.20 and 89.97%±1.30, respectively (p< .05). Based on the in vivo test, pretreatment with 2 mg/kg of colchicine one hour before tachyzoites injection had the best inhibitory effect (70.32%±4.07). Also, pretreatment with 2 mg/kg of propranolol 90 min before tachyzoites injection (78.54%±1.99) induced the best inhibitory effect (p< .05).

According to the results, colchicine and propranolol could inhibit tachyzoites entrance into nucleated cells in vitro and in vivo. In this study, the most efficient concentrations and times for using these substances were determined.

According to the Iranian Traditional Medicine (ITM) references, Platanus orientalis L. possesses wound healing properties. Herein, we developed different topical formulations based on the ethanolic extract of P. orientalis leaves and evaluated its wound healing effects through an in vivo model.

Hydroalcoholic extract of the leaves was obtained from ethanol 80% and it was evaluated for DPPH radical scavenging activity, total phenolic and flavonoid contents as well as the presence of tannins. Different topical formulations including ointment (D-O) and polymer film (D-F), were prepared and an in vivo test was run for 14 days in an excision wound model consisting of 5 groups of 6 rats.

The results indicated the higher efficacy of D-O compared with D-F, as wound surface area remarkably reduced within 14 days post-injury. Also, histological features including epitheliogenesis score, neovascularization, and collagen density indicated the potential wound healing effect of D-O.

Wound healing properties of the ethanolic extract of P. orientalis leaves depended on the type of formulation and D-O was found to be much more potent than D-F, from reducing wound surface area, maximum epitheliogenesis score, proper neovascularization pattern, and early type I collagenization points of view.

In the present study, we investigated the application of pulsed magnetic field (MF) (3.5 T, 1 Hz, 8 square-wave/160 µs) permeabilization on murine breast adenocarcinoma cells when administering bleomycin (BLM) in vivo.

This cross-over study aims to find a noninvasive method to facilitate penetration of hydrophilic anti-cancer drugs through the cancerous cells membrane into the cytosoll in order to minimize the side effects of the chemotherapy treatments of tumors.

In this cross-over study, a total of 50 female Balb/c mice were tumorized via homograft. After about 2 weeks, magnetic pulses (3.5 T, 1 Hz, 8 square-wave/160 µs) were applied to tumor-bearing mice 3 min after intratumoral BLM solution injection. Tumor volume was measured every 48 h during 22 days.

The results showed that the difference between the BLM plus 3.5 T MF group versus the sham control or sham MF groups was significant. Uptake of BLM molecules by tumoral cells in the BLM plus 3.5 T MF group versus the BLM control group was 7- folds higher that this result was statistically insignificant (p <0.05, SEM=266.8676, analysis of variance).

Significant cell permeabilization to BLM requires greater MF strength or exposure time. Further investigation is necessary.

Benzodiazepines (BZD) are among the main classes of tranquilizing drugs, bearing much less toxicity compared to other drugs acting on the CNS. Considering the pharmacophore model of BZD binding to GABA-A receptor, novel diphenyl 1,3,4-oxadiazole compounds as BZD ligands were designed. The compounds were synthesized and structurally confirmed using LCMS, IR and NMR techniques. We investigated the affinity of the compounds to BZD receptors using radioligand [3H]-flumazenil by in-vitro studies. In addition, sedative-hypnotic, anxiety, anticonvulsant, muscle relaxant, memory impairment, and motor coordination activities of the synthesized compounds were evaluated using in-vivo studies. Based on in-vitro studies, compounds 7i and 7j were the most potent with IC50 values of 1.54 and 1.66 nM respectively. In-vivo studies showed that compound 7i has the highest impact on increased sedation, muscle relaxation, and decreased anxiety and these observations were antagonized by flumazenil. Compounds 7e and 7i were the most potent anticonvulsant agents among synthesized compounds in both MES and PTZ induced seizure tests. All synthesized compounds significantly decreased latency to fall in the Rotarod test but none of them had a significant impact on the memory impairment test.