Therapeutic potential of Chemogenic Iron oxide Nanoparticles, its antioxidant and anti-Inflammatory effects with reduction of ankle joint swelling in BALB/c mice

Data availability

Data will be made available on request.

The data that support the findings of this study are available from the corresponding author upon reasonable request.

1 Introduction

Nanoparticles are extraordinary small size particles with size ranges between 1 and 100 nm either in two or three dimensional orientation (Oborny, 2007). This unique nature of the nanoparticles are incredibly useful in drug delivery (Mudshinge, Deore, Patil, & Bhalgat, 2011). Having said of their nano size, they can easily approaches the targeted location within the human body and bypass the biological barriers. Nanoparticles can be engineered for transportation and to prevent the degradation of drugs and therapeutic substances and it also allow slow and control release of these loaded substances. Moreover the surface of nanoparticles can also be modified for target drug delivery particularly to tissues and cells. The improved and enhance target drug delivery lowers the risk of drugs side effects (A. Kumar & Thakur, 2022).

Iron Oxide nanoparticles consisting of large variety of polymorphs in nanomaterials with unique physiochemical characteristics. They may have 4 standardize principal polymorphic forms, hematite (α-Fe2O3), β-Fe2O3, maghemite (γ-Fe2O3) and ε-Fe2O3; similarly the iron (III) oxy-hydroxide also comprises four polymorphic forms which are recognize as goethite, akaganeite, lepidocrocite and feroxyhyte as α, β, γ and δ-FeOOH respectively. Interestingly, α-Fe2O3 is a well-known n-type semiconductor having bandgap Eg = 2.1 eV, prominent for an excellent chemical stability and reduce toxicity under normal existing conditions. Out of theses mentioned oxides, α-FeOOH and α-Fe2O3 have been popular for magnetic devices, catalysts, gas sensors, effective UV-blocking (Havenga et al., 2022) and photoelectrodes applications (Matinise et al., 2018). Moreover the green synthesized iron oxide nanoparticles has gain popularity due to its ecological friendly approach and improved the biocompatiability (Hassan et al., 2018; Khalil et al., 2017; Köçkar, Karaagac, Özel, & Materials, 2019), antimicrobial activity (Aisida et al., 2020; Madubuonu et al., 2020).

Gout, a painful joint inflammation, affects approximately 1% of the population, predominantly males (Wu et al., 2022). It occurs because monosodium urate crystals (MSUC) are gathered in the joints and surrounding tissues. Gout patients often exhibit hyperuricemia, characterized by raised uric acid levels in the blood (>70 lg/ml). Acute gout attacks can be triggered by factors like injury (Abhishek, Valdes, Jenkins, Zhang, & Doherty, 2017), exposure to cold, high food intake, and obesity (Tan, Farrar, Gaucher, Miner, & evolution, 2016). Uric acid is a necessary intermediate in the breakdown of purines, and its serum levels can rise due to various factors, including genetic factors and lifestyle choices. In most organisms, uric acid is transformed into allantoin, a soluble material, by the enzyme uricase. However, in primates, including humans, uricase activity is diminished due to gene mutation (Parthasarathy, Vivekanandan, & systems, 2018), resulting in the accumulation of uric acid in the body. Uric acid has the lowest solubility among purine byproducts, and its crystallization into MSUC is a critical factor in uric acid-mediated inflammation seen in gout. The inflammatory response stimulated by MSUC can lead to the development of tophi, which accumulates urate crystals in the tissues and joints (Kosugi, Nakagawa, Kamath, & Johnson, 2009). Some studies have evident that raised levels of soluble uric acid may be linked with conditions such as hypertension and certain cardiovascular diseases (S. Kumar et al., 2019).

The broad spectrum utilization of iron oxide nanoparticles within different applied scientific fields (O Karaagac, Hasirci, & Köçkar, 2024), such as controlled drug delivery systems, enzyme immobilization, and medical diagnostics, is more interesting area of research. Iron oxide nanoparticles demonstrate remarkable qualities in medicine with their exceptional characteristics like high surface area, magnetic susceptibility, and biocompatibility (Bhatia, Bhatia, & algae, 2016). The combine impact of nanotechnology and drug delivery system holds great prospective for improving the cure of enormous diseases, including those related to bone health and uric acid metabolism (Yang & Webster, 2009). Ongoing research and developmental efforts in this field have potential to uncover cutting-edge and unique therapeutics that can improve patient outcomes and quality of life (Oznur Karaagac, Kockar, Tanrisever, & magnetism, 2011; Oznur Karaagac, Kockar, & magnetism, 2012).

In this current study, researchers have synthesized iron nanoparticles to investigate their effects on the reduction of ankle diameter, antioxidant and anti-gout properties in a gouty mice model.

2 Material and methods

2.1

2.1 Synthesis of Iron oxide nanoparticles

The iron oxide nanoparticles was obtained using the Sol-Gel precipitation method. A 1:2 ratio of the two iron salts FeCl₂ and FeCl₃ were dissolved in continuous stirring 50 ml of deionized water and then 50 ml of 8.5 M NaOH solution were added drop by drop. When precipitates were formed then the resulting mixture were undergone the filtration process, the precipitates obtained were then washed multiple time with deionized water. After the complete washing the precipitate is then subjected to calcination at 100°C for overnight which was the main step for iron oxide nanoparticles formation. Powder form Iron oxide nanoparticles were obtained after overnight calcination which were grinded using a mortar and pestle. This whole process of Iron oxide nanoparticles synthesis is represented in Fig. 1 with slight modifications of previously conducted study by Karaagac and Kockar (Oznur Karaagac et al., 2012).

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Fig. 1

Graphical abstract; (A) Synthesis and characterization of iron oxide nanoparticles, (B) Induction of gout, (C) Treatment of mice with iron oxide nanoparticles, (D) Results.

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2.6

2.6 Experimental design

The MSUC induced hyperuricemia experimental mice were divided into six groups, all receiving MSUC, except for the control group. MSUC were injected intra-articularly in left ankle and intra-peritoneal over three weeks and two weeks respectively at one-day intervals. Blood samples were evacuated regularly to monitor the mice's uric acid concentration. After the initial three-week period, the mice were subjected to oral administration of various concentrations of iron oxide nanoparticles, as specified in Fig. 2. Upon completion of the experiment, the animal model mice were euthanized; blood and tissue samples of Kidney, liver, and muscle were collected and preserved within paraffin blocks for subsequent analysis of tissue iron levels using atomic absorption spectroscopy and for histopathology determination (Kakkar, Das, & Viswanathan, 1984).

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Fig. 2

Schematic diagram presenting dose administration in MSUC induced gouty mice.

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3 Results

3.1

3.1 Characterization of nanoparticles

The X-ray diffraction analysis of the produced iron oxide nanoparticles corresponds to hematite indicates several Bragg reflections at different Bragg angles, as demonstrated in Fig. 3. Hematite exhibits a rhombohedral crystal structure with Fe³⁺ ions occupying octahedral coordination sites in a hexagonal close-packed array of oxygen ions. According to the previous study (Kiyani, Moghul, Butt, et al., 2022) the intense peak observed at around 2θ of 35.6748 corresponds to the (104) plane, characteristic of hematite α-Fe₂O₃ as per JCPDS card no. 33–0664. Moreover additional peaks further support its identification and is aligned with the electronic valence of Fe³⁺, which confirms trivalent oxidation state of iron within the hematite phase. The estimated particle size was to be 58.64 nm by using the Debye-Scherrer equation. The SEM images showed morphological spherical shape with minimal agglomeration of synthesized iron oxide nanoparticles with approximately 64 nm in size as depicted in Fig. 4 (a) Moreover, energy-dispersive X-ray (EDS) established the occurrence of iron in the desired quantity, along with traces of other elements like chloride and sodium, EDS showcases in Fig. 4 (b). The magnetic hysteresis curve for the iron oxide nanoparticle sample is given in Fig. 5. The results presented in the measurements show weak dependence on magnetic fields and negligible hysteresis at low magnetic fields. The saturation magnetization M_s was resolved to be approximately 0.700 emu g⁻¹. H_c covering the magnetic field necessary to reduce the magnetization to zero after saturation was calculated to be −40.08 Oe. These results indicate the assumption made here that the nanoparticles are superparamagnetic as indicated by the fact that they have very low coercivity and zero remanent magnetization.

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Fig. 3

X-ray diffraction pattern (XRD) of synthesized Iron oxide nanoparticles.

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Fig. 4

(a) Scanning Electron Microscope (SEM) image of synthesized Iron oxide nanoparticles (b) Energy dispersive X-ray spectroscopy pattern of synthesized Iron oxide nanoparticles.

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Fig. 5

Hysterisis Plot for SQUID analysis showing M vs. H for iron oxide nanoparticle at 300 and 4 K.

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4 Discussion

The findings of the current study demonstrated that Iron oxide nanoparticles show promise in the treatment of gout in mice resulting from hyperuricemia, even at lower concentrations, while minimizing excessive damage to cells. Additionally, various biochemical tests were performed in this research work to assess the impact of Iron oxide nanoparticles on mice. Dextran or starch is commonly employed as a stabilizing agent in forming iron oxide nanoparticles to create a system that can disperse in water (LaConte, Nitin, & Bao, 2005). In an earlier study, the impact of injected Iron oxide nanoparticles on oxidative stress in various tissues and monitoring changes in hepatic enzymes as a measuring parameter of liver function was assessed. During investigations, a gradual rise in serum iron levels over one week was followed by a slow drop down, indicating that the elimination of magnetic NPs from the body takes longer than three weeks. The increase in serum iron levels can be attributed to several concurrent processes, including the breakdown of injected iron particles, subsequent binding of iron to various type of serum proteins like transferrin and re-dispersion of bound iron to body cells and tissues, and eventual elimination. The study suggests that most body iron exists in the bound state due to its slow clearing kinetics (Jain, Reddy, Morales, Leslie-Pelecky, & Labhasetwar, 2008).

One of the previous research observed that administration of silver oxide nanoparticles resulted in a notable elevation in protein estimation and lipid profile values in a group of gouty mice induced by MSUC. The raised protein levels indicated the magnitude of the response of inflammation, which is closely linked to oxidative stress (Kiyani, Moghul, Javed, et al., 2022). A separate study concluded that administration of ZnO-NPs to hyperuricemia mice model with gouty arthritis resulted in a considerable reduction in levels of blood uric acid. Moreover, the lipid profile tests demonstrated that the treatment with nanoparticles effectively lowered cholesterol and LDL levels with comparison to the control group. However, both the nanoparticles-treated and allopurinol-treated groups exhibited increased in hepatic enzymes concentrations.

Nonetheless, histopathological examination of kidney, liver and muscle tissues revealed no noticeable variations. This research provides evidence supporting the potential of orally given iron oxide nanoparticles in reducing the concentration of uric acid, cholesterol as well as LDL of the blood (Kiyani et al., 2019). In a previous conducted research, it was reported that CuO nanoparticle administration at different concentrations reduced serum ankle swelling in gouty mice with comparison to the negative control group (Kiyani et al., 2020), and these results are similar to our findings that Iron oxide nanoparticles administration reduced swelling in all treated groups.

5 Conclusion

This research results represents that low concentrations of Iron oxide nanoparticles hold potential as a prospective remedy for gouty arthritis in mice. Administering gout-afflicted mice with Iron oxide nanoparticles at 125, 250 and 500 mg/kg of body weight showcased antioxidative effects, signifying their possible therapeutic application in gout treatment. Moreover, Iron oxide nanoparticles demonstrated the capacity to lower total protein levels, suggesting reduced inflammation associated with gout.

CRediT authorship contribution statement

Mubin Mustafa Kiyani: Writing – original draft, Methodology, Conceptualization. Sarah Sadiq: Formal analysis, Data curation, Conceptualization. Maria Sarfraz: Investigation, Funding acquisition, Formal analysis. Chanda Javed: Resources, Methodology, Investigation. Maisra Azhar Butt: Software, Resources, Project administration. Hassan Burair Abbas: Project administration, Investigation, Funding acquisition. Hamza Rehman: Validation, Software, Project administration. Syed Ali Imran Bokhari: Writing – review & editing, Visualization, Validation, Supervision.

Funding

This research was conducted without any external funding.