174 

© 2024 The Author(s). Published by College of Education for Pure Science (Ibn Al-Haitham),             

University of  Baghdad. This is an open-access article distributed under the terms of the Creative Commons 

Attribution 4.0 International License 

 

 

Chitosan Reinforced Polyvinyl Alcohol and Nanoparticles Treatment to 

Promote Skin Wound Healing in Diabetes Induced Mouse Model 
 

Huda R. Kttafah1,* , Hanaa S. Mahmood2  and Hussein N. Olewi3  

 
1Department of Physics, Ministry of Education, Directorate General of Education Rusafa 2, Baghdad, 

Iraq. 
  2Department of Physics, College of Education for Pure Science, (Ibn Al-Haitham), University of 

Baghdad, Baghdad, Iraq. 
3Department of Mechanical Engineering, College of Mechanical Engineering, University of Karabuk, 

Karabuk, Türkiye. 
*Corresponding Author. 

 
Received: 2 November 2023                   Accepted: 8 January 2024                  Published: 20 October 2024 

doi.org/10.30536.37.4.3808 

 

Abstract   

A group of mice were captured. We divided the 3-month-old mice into groups. We limited 

each group to a specific type of treatment, using different components of silver (Ag) and gold (Au) 

in various proportions. The control group was left untreated. We stimulated her diabetes with 

injections of the substance alloxan, which led to the development of her condition. It was left for 

three days after the injection of this substance, and then the diabetes was checked with a diabetes 

testing device until all the required mice were infected with diabetes. Next, they removed the hair 

from the mouse's top and used a special tool, a ring with a diameter of 8 mm, to cut the skin. During 

the work, they used two types of anesthetics: ketamine and xylazine. The ketamine relaxes her 

muscles and causes her death, so she abandoned the xylazine and relied solely on the first type. 

The percentage of the anesthetic substance injected depends on its weight. Therefore, we created 

an incision on the mouse. Then I began treating the wounds: the first group with 100% AgNPs, 

the second group with 100% AuNPs, and the third group with samples of a mixture of AgNPs and 

AuNPs at a ratio of 50% each. The fourth and final group remained without treatment. With a 

weekly checkup for diabetes. The treatment entailed applying the sample to the wound and 

adhering it with a medical adhesive, periodically replacing it. After the second week of treatment, 

some of the wounds began to heal. By the end of the fourth week, it had completely healed. The 

group that was treated with the mixed samples had the best and fastest recovery and wound healing 

among the rest of the samples, followed by the mixture samples. 

Keywords: Chitosan/PVA, nanoparticles, skin healing, diabetes. 

 

1. Introduction 

Skin physiological impairments in uncontrolled diabetes mellitus (DM) patients are 

frequently associated with delayed and challenging healing processes. In an animal-induced DM 

model, mimicking the wound phenotype would facilitate the assessment of wound-healing 

products and the healing duration period. We looked at how well chitosan/PVA-embedded Au and 

Ag nanoparticles helped wounds heal faster in a mouse model than alloxan-induced DM. We need 

modest and appropriate animal models in wound healing, and these trends are welcome in human 

https://creativecommons.org/licenses/by/4.0/
https://creativecommons.org/licenses/by/4.0/
https://orcid.org/0009-0008-9934-1572
mailto:Huda.Razaq2104p@ihcoedu.uobaghdad.edu.iq
https://orcid.org/0009-0002-3755-1298
mailto:Hanaamahmood43@yahoo.com
https://orcid.org/
mailto:hussinasr6@gmail.com


IHJPAS. 2024, 37 (4)  

175 
 

wound healing. Therefore, today's wound treatment is more effective and better understood than 

in the past. We aim to develop the most suitable animal model for human use, which also 

contributes to the healing and treatment of wounds. 

Diabetes is chronic for many people who suffer from a deficiency in insulin secretion, which in 

turn leads to high blood sugar [1]. Diabetes is one of the most pressing global health issues in all 

countries. In the coming years, we expect the number of affected individuals to increase [2]. 

Diabetes is heat-resistant and affects all organs and tissues of the body, including the skin. A 

diabetic person faces the problem of faster wound healing compared to non-diabetic individuals 

[3]. Many diabetics suffer from skin problems, and this is obvious to them. They will easily 

develop skin diseases as they grow older [4]. It may lead to skin deformities associated with stress 

and pressure, poor growth factor production, poor blood vessel formation, and abnormal regulation 

of inflammatory products, all of which contribute to premature and accelerated skin aging [5]. It 

also contributes to the progression of skin infections. The inflammation may reach the deep layer 

of the skin and cause serious damage, which in turn is a medical burden [6]. 

Indeed, this is a significant warning sign for individuals with diabetes. The non-healing of the 

wound is attributed to a decrease in the production of fats in the skin, leading to dryness, and a 

decrease in the production of platelets, which are responsible for blood clotting in wounds [7]. It 

causes bleeding and increased inflammation. This may lead to the amputation of the affected part 

[8]. The environment in which the injured person lives plays a fundamental role in the patient’s 

recovery, in addition to the new treatment. [9]. 

In this research paper, I investigated the effects of diabetes on the skin and how to treat wounds. 

The study found that diabetes accelerates the healing process in mice compared to non-infected 

mice [10]. Understanding the primary causes of skin complications in diabetic patients could aid 

in early diagnosis and treatment planning [11]. It has the potential to benefit humans and aid in 

their recovery from diabetes [12]. We utilized both natural and synthetic polymeric materials, 

including chitosan, chitosan, and polyvinyl alcohol. Chitosan was the basic material. We used 

prepared nanoparticles as solutions to boost the insulin levels of the diabetes-infected mice in this 

study, which consequently reduced their blood sugar levels [13]. 

 

2.  Materials and Methods 

We prepared the matrix, which is the chitosan powder, by mixing it with acetic acid and 

obtaining a homogeneous solution. Next, we prepared the reinforced material, powdered polyvinyl 

alcohol, by mixing it with distilled water. We then mixed the two solutions to create a single, 

homogeneous solution. Two types of nanoparticles, prepared in different proportions of both silver 

and gold, reinforced the resulting solution. Consequently, we acquired nanoscale samples to 

manage diabetic wound aggregates. Study design: experimental groups. The three treatments and 

the control group consisted of 4 BALB-c mice each (Tab the University of Baghdad College of 

Veterinary Medicine in Abu Ghraib, Iraq provided the BALB-c mice from their animal laboratory. 

We acclimatized the mice in special cages for two weeks, maintaining a temperature of 24 °C, 

medium humidity, and a light and dark cycle of 12–12 hours. ours. Access to drinking water and 

food was exceedingly easy [13]. 

 
Table 1. Experimental groups. 

  Treatment groups Control 

Group # G1 G2 G3 G4 

# of mice 4 4 4 4 

Treatment 100 % Ag NPs 100%AuNPs (50%Ag+50%AuNPs) no treatment 

 

 

 

 



IHJPAS. 2024, 37 (4)  

176 
 

In BALB-c mice, alloxan-induced diabetes occurs. After a 12-hour fast, all four groups were given one 

or more intraperitoneal (IP) injections of alloxan (Serial No. AL1027B, 2-Alloxan (hydrate) 98%, 

Batch No. Aoo89B) by ALPHA CHEMIKA at a dose of 100 mg/kg of body weight, as explained in 

[14]. A diabetes-measuring device was used to collect a blood sample from the tail after three days of 

alloxan administration, we regularly tested the blood glucose single-point levels to confirm the 

induction of diabetes; the fasting glucometer readings were > 250 mg/dL. mg/dL. We dissolve the 

alloxan (100 mg powder) in 10 ml of distilled water and calculate the dosage as follows [15]. 

Dosage in mg = 
𝐵𝑜𝑑𝑦 𝑊𝑒𝑖𝑔ℎ𝑡 𝑜𝑓 𝑎𝑛𝑖𝑚𝑎𝑙 (𝑔 )

1000 𝑔
∗ 𝑑𝑜𝑠𝑒(𝑚𝑔)                                                                      (1) 

We performed the wound generation on the back side of the mice's bodies in all four groups of animals 

Figure 1 [16]. We sedated the animals with 0.1 mg/kg of BW ketamine (subcutaneous injection) and 

excised a skin circle-shaped punch (8 mm diameter) from the back of the mice. We applied the 

treatment using a wound dressing bandage, adhering it to the entire excision area and replacing it with 

a new one every week [17]. Every week, we take a skin biopsy to replace the dressing bandage 

treatment. We removed the initial skin, saved all subsequent biopsies in a buffer, and sent them for 

histopathological examination.[19]. 

 

Figure 1. Skin wound generation and treatment application. Skin wound excision (circle shape 8mm in diameter) (left 

picture). Wound treatment using dressing bandages contains the Au, Ag NPs combination treatments as explained in 

the experimental group's Table 1. (right picture).  

 

3.  Results and Discussion 

To characterize the tissues of the cells of the epidermis and dermis, we use the electron 

microscope, which is considered a complement to the histological analysis. Through the 

microscope, it is clear that the animal's skin structure is a difference in the structure of the skin of 

the animal. The organization of keratinocytes in mouse strains resembles a honeycomb. There 

were no microscopic pathological changes in the skin of the animals taken for both the negative 

control group and the animals of the other groups before treatment. As shown below in Figure 2 

[20], the epidermis and dermis compose the skin, characterized by their typical layers without the 

appearance of any lesions. 

 



IHJPAS. 2024, 37 (4)  

177 
 

 
Figure 2. Histopathological figures for the section of skin for one animal in the control the negative group showed 

epidermis and dermis with no legion (H&E(X20)). 

 

The control group, the positive group, showed thickening of the epidermis (layers) in all its layers, 

especially the stratum spinosum [21]. The cells showed necrosis (the presence of gaps) in the layers 

of the epidermis as well as proliferation of fibroblasts and generators, as shown in Figure 3 [22]. 

 

 
Figure 3. The histopathological figure for |section of skin for one animal in the control the Positive group showing 

thin new of epidermis vacuolation of call-in epidermis (A) proliferation of fibroblast (B) (H&E(X200)). 

 

In addition to the infiltration of inflammatory cells in the epidermis and dermis layers and around 

the hair follicles, as shown in Figure 4[23]. 

 

B 
A 



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178 
 

 
Figure 4. The histopathological figure for the section in skin for one animal in the control the Positive group induced 

by D.M shows thin new infiltration of the inflammatory cell (A) in addition to peri hair follicles accounting epidermis 

(B) and of number Phylis (C)(H&E(X200)). 

 

Most of these cells are neutrophils[24]. It even collected neutrophil cells in some sections and also 

cleansed the abscess, as shown in Figure 5. 

 

 
Figure 5. The histopathological figure for the section of skin for one animal in the control the Positive group showed 

abscessation of hair follicles (A) with the proliferation of fibroblast (B) and infiltration of inflammatory cells (C) 

(H&E(X400). 

 

The histopathological changes in the group treated with silver nanoparticles showed fewer 

pathological changes than those found in the positive control group [25]. There were fewer 

thickenings than in the positive control, as shown in Figure 6. In addition to the presence of 

hyperplasia of hair follicles and fat cells, along with the proliferation of fibroblasts and 

fibroblasts[26]. 

C 

B 

A 

C 

A 

B 



IHJPAS. 2024, 37 (4)  

179 
 

 
Figure 6. The histopathological figure for a section of skin for one animal in the 100% AgNPs treated group shows 

(A) Epidermis skin layer thickening and fibroblast proliferation. (B) hyperplasia of the hair follicles and the sebaceous 

glands. (H&E-stained skin lesion X40). 
 

If you looked closely at the tissue changes in the group that was given gold nanoparticles, you 

could see that they had a net of fibrous connective tissues, inflammatory cells invading, and a lot 

of fibroblast cells and sebaceous glands rising [27]. Moreover, the positive pathological changes 

in the positive control group were more severe than those in the gold nanoparticle-treated group 

[28]. The severity of the inflammation in the gold group caused necrotic areas to appear in some 

sections, but they were minor [29]. 
 

 
Figure 7. The histopathological figure for section Shows one animal treated with 100% AuNPs infiltration of the 
inflammatory cell(A) with fibrous connective tissues (B) and sebaceous glands (C)(H&E(X20)). 

 

 
 

 

 

 

 

 

 

 

 

 

 

 

 

Figure 8. The histopathological figure for section Shows for one animal treated with 100% AuNPs Thickening of 

the epidermis (A), infiltration with inflammatory cells(B), and necrotizing of sebaceous glands (C) (H&E-stained 

skin lesion (X40)). 

 

B 
A 

C 

B 

A 

A 

A 

A 

B 

C 



IHJPAS. 2024, 37 (4)  

180 
 

 
Figure 9. The histopathological figure for section Shows one animal treated with 100% AuNPs proliferation of the 

hair follicles (A), infiltration of inflammatory cells (B), and proliferation of fibroblast cells(C) (H&E-stained skin 

lesion (X200)).  

 

The mixture group containing 50% AgNPs and 50% AuNPs exhibited very slight pathological 

changes. The treatment resulted in a noticeable and obvious improvement, as evidenced by the 

reduced thickening of the epidermis layer and the absence of inflammatory cell infiltration [30]. 

Noting that hair follicles and seborrheic cells were naturally present, even some sections were 

closer to normal (natural tissue), as shown in Figure 10 and Figure 11 [31]. concurring with 

Maycon Carvalho Ribeiro's research, which applied nanoparticles and chitosan to treat diabetic 

patients' wounds more quickly in rats. Also agreed with the researcher Paola Losi, who created an 

application to speed up wound healing in diabetes [32]. 
 

 

 

 

 

 

 

 

 

 

 

 

 

 
 

 

 

Figure 10. The histopathological figure for section Shows one animal treated with 50% AgNPs and 50% AuNPs mix 

treated group (A) with a slight thickening of the epidermis with few numbers of fibroblast (H&E-stained skin lesions 

(X10)). (B) normal hair follicles and the sebaceous glands (H&E-stained skin lesion (X40)). 

 

B 

A 

C 

A B 



IHJPAS. 2024, 37 (4)  

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Figure 11. The histopathological figure for the section Shows that one animal treated with 50% AgNPs and 50% 

AuNPs mix treated group connective tissue is closer to natural (H&E-stained skin lesion (X40)). 
 

This study found that there was a difference in response to treatment with nanoparticles among all 

groups. The group that received samples supported by a mixture of gold and silver nanoparticles 

in equal proportions achieved the best results. The group that received samples supported by gold 

nanoparticles came next. When the group was treated with silver nanoparticles, it responded less 

to treatment. The control group had skin infections, and recovery was very slow. 

  

5. Conclusion 

         From the current study and through the use of nanoparticles in the treatment of diabetes, I 

found that using a mixture of silver and gold nanoparticles at a ratio of 50% of each has the greatest 

role in treating the wounds of diabetic patients, in addition to speeding up healing. This mixture is 

better than using each type of nanoparticle alone. 

 

Acknowledgment  

        I would like to thank the staff of the Physics Department, Deanship of the College of 

Education for Pure Science (Ibn Al-Haitham) for their support in writing this research.  

 

Conflict of Interest 

        The authors declare that they have no conflicts of interest. 

 

Funding 

         None. 

 

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