To study the effect of activated platelet-rich plasma in cases of androgenetic alopecia

Vijay Kumar; Navneet Sharma; Brijesh Mishra; Divya Upadhyaya; Arun Kumar Singh

doi:10.4103/tjps.tjps_86_20

ORIGINAL ARTICLE

Year : 2021 | Volume : 29 | Issue : 5 | Page : 28-32

To study the effect of activated platelet-rich plasma in cases of androgenetic alopecia

Vijay Kumar¹, Navneet Sharma², Brijesh Mishra³, Divya Upadhyaya³, Arun Kumar Singh⁴
¹ Rajarshi Dashrath Autonomous, State Medical College, Ayodhya, U.P, India
² AIIMS, Bilaspur, H.P, India
³ KGMU, Lucknow, U.P, India
⁴ Atal Bihari Vajpayee Medical University, U.P, India

Date of Submission 29-Jul-2020

Date of Acceptance 27-Oct-2020

Date of Web Publication 17-Mar-2021

Correspondence Address:
Dr. Navneet Sharma
Assistant Professor, Department of Burns and Plastic Surgery, AIIMS, Bilaspur, Himachal Pradesh
India

Source of Support: None, Conflict of Interest: None

DOI: 10.4103/tjps.tjps_86_20

Abstract

Context: Growth factors released from platelets act on stem cells in the bulge region of follicles and thus stimulate the development of new hair follicles. Aims: Primary aim of the study was to assess the efficacy and safety of activated platelet rich plasma in AGA. Settings and Design: Prospective double arm non-randomized pre-post study. Material and Methods: Activated Platelet-rich plasma was injected in either anterior half of alopecia scalp in first 20 patients and posterior half in next 20 patients (Group I) and the other half was injected with placebo (Group II). Each patient received a total of 5 session of aPRP, 15 days apart. Statistical analysis used: Quantitative variables were compared using student T test and Mann-Whitney Test between two groups. A p value of < 0.05 was considered statistically significant. Results: 40 male patients with a clinical diagnosis of AGA (stage II to VI) were recruited but only 35 patients completed the whole study. At 1 month and 3 months follow-up, Group I showed statistically significant increase in hair count, hair density and anagen/telogen ratio as compared to Group II. Conclusion: aPRP is a safe and good alternative to medical treatment. It seems to have a stabilizing effect on hair fall and may be considered as a measure to prevent progression of alopecia in early stages.

Keywords: Activated platelet-rich plasma, androgenic alopecia, growth factors

How to cite this article:
Kumar V, Sharma N, Mishra B, Upadhyaya D, Singh AK. To study the effect of activated platelet-rich plasma in cases of androgenetic alopecia. Turk J Plast Surg 2021;29, Suppl S1:28-32

How to cite this URL:
Kumar V, Sharma N, Mishra B, Upadhyaya D, Singh AK. To study the effect of activated platelet-rich plasma in cases of androgenetic alopecia. Turk J Plast Surg [serial online] 2021 [cited 2023 May 30];29, Suppl S1:28-32. Available from: http://www.turkjplastsurg.org/text.asp?2021/29/5/28/311440

Introduction

Androgenic alopecia (AGA) is a type of nonscarring alopecia which is characterized by progressive diminution in size of the hair follicles. It is a common problem which can affect both males and females. Affected individual may see this male pattern baldness with shame and resultant injurious consequences such as low self-perception.^[1],[2]

It is an androgen-dependent disorder which is mediated through interaction between dihydrotestosterone (DHT) and the hair follicle-related androgen receptor (AR) which results in gradual shortening of anagen phase and lengthening of telogen phase. As a result, length and diameter of affected hair decrease, thus leading to bald appearance.^[3] Minoxidil, 5-alpha-reductase inhibitors, and hair transplantation are most commonly used treatment modalities with curative intent but with variable success rate. All these modalities have their own limitations and side effect profile.^[4]

To overcome these limitations, cell-based therapies and tissue engineering are of heightened interest in the present era. Platelet-rich plasma (PRP) is one of the emerging modalities. It is hypothesized that growth factors released from alpha granules, enhance vascularity, and new hair growth by acting at stem cells in bulge region of the hair follicles.^[5] To release these growth factors, platelets need to be activated, which can be done by adding thrombin or calcium chloride in freshly prepared PRP.^[6] Role of PRP for the treatment of male pattern hair loss has also been successfully demonstrated in many studies, but there are very few studies which have used activated PRP (aPRP).^[7],[8],[9] The aim of this study is to demonstrate the effectiveness of aPRP in AGA.

Methodology

This was a prospective double arm non-randomized pre-post study conducted on forty patients with AGA. This study was carried out after getting approval from the institutional ethical committee wide reference no ECR/262/Inst/UP/2013/RR-16/Protocol No: 859/Ethics/R. Cell-18 at KGMU, Lucknow, between June 2018 and May 2019. Male patients aged between 18 and 50 years with a diagnosis of male pattern hair loss (Grade II–Grade VI, according to Hamilton-Norwood Classification), with no previous medical, surgical, or psychiatric ailment and who had given written consent were finally included in this study. Patients who had taken medical treatment for AGA within the past 6 months were excluded from the study.

The efficacy and safety of aPRP was assessed in all patients of AGA. Phototrichograms (hair count [hair/0.65 cm²], hair density [number of hair/cm²], hair diameter, anagen/telogen hair ratio, and terminal/vellus hair ratio) were compared at baseline, 1 month, and 3 months follow-up in Group I (aPRP intervention group) and Group II (placebo group).

Approximately, 20 mL of blood was drawn into a syringe prefilled with 2 ml of 3.8% sodium citrate anticoagulant. Centrifugation was performed at 2000 rotation per minute (rpm) for 5 min (Eppendorf centrifuge 5804/5804R). The sample was separated into three layers – upper yellowish plasma, a thin middle buffy coat layer of platelets and white blood cells, and a red bottom layer of RBCs. The upper two layers were transferred to an empty sterile tube, and second centrifugation was performed at the force of 2500 rpm for 10 min. Upper two-third of fluid was removed, and the rest left out was homogenized. PRP (approximately 2–3 ml) was reconstituted with 10% calcium chloride (approximately 0.2–0.3 ml) in a ratio of 9:1 to form aPRP, just before injecting into the scalp.^[10]

PRP was injected in the anterior half of alopecia area in the first twenty patients, and in posterior half of alopecia area in next twenty patients, while normal saline was injected in other half of all patients with insulin syringe. A paper template with a round hole in the center, of approximately 1 cm, was placed on the scalp surface at the center of each half, and clippings of hair in these two target areas were performed, and digital images (Phototrichograms) were performed in all patients by means of Cooling Tech microscope, and digital image analysis was done. Cooling Tech microscope is a digital software-supported system which took images with a handheld microscope and all hair parameters were analyzed with the help of software. A semi-permanent dot tattoo was placed in each of these target areas, which guarantees the analysis of the same area and therefore ensures the reproducibility of the study. The tattoo was visible during all study periods. Forty-eight hours after clipping, images of the selected areas were again obtained. The digital images were standardized, always at the same distance from the scalp surface and with a 20-fold magnification. All recorded phototrichograms were manually analyzed using the Cooling Tech microscope software. The hair parameters were obtained for each image. The infiltration aPRP and normal saline were carried out with a 30G needle at a dermal level in a linear fashion at depth of 2–2.5 mm and 1 cm apart.

All patients were observed for at least 1 hour after procedure for any complication. The procedure was repeated every 2 weekly for five sessions with injection of PRP and normal saline in the same field as allotted in the first sitting.

The patients were photographed at the time of enrollment, at 1-month and 3-month follow-up taking all standard views (frontal, lateral, vertex, and back view). All photographs were taken in standardized illumination and background using same digital camera. The photographs taken before, during, and after the treatment were compared and assessed by assessor blinded from the study. The qualitative changes were graded using a hair profile scale. Hair profile scale is comprised of five grades, i.e., dcreased hair, unchanged, mild improvement, moderate improvement, and marked improvement.

Subjective improvement of patients was assessed with the help of a questionnaire. The questionnaire contained five entities: (1) change in hair quality, (2) change in hair volume, (3) decrease in shedding of hair, and (4) pigmentary changes and hair regrowth. Patients were asked to rate them at end of treatment and response was graded as: no change (−), slight change (+), moderate change (++), and significant change (+++) depending on response to treatment.

Data analysis

Normality of data was tested by Kolmogorov–Smirnov test. If the normality was rejected, then nonparametric test was used. Quantitative variables were compared using Student's t-test and Mann–Whitney test between two groups. A P < 0.05 was considered statistically significant.

Results

Forty patients of AGA were recruited, of which only 35 patients completed the whole study and five patients did not comply with the study protocol and left in between the study. The study samples were between 20 and 50 years of age, with a mean age of 26.16 ± 9.88 years and the median age was 27 years. Clinically, and according to Hamilton–Norwood Classification, the most frequent pattern of baldness in our study was Stage V (40%), followed by Stage IV (37%), Stage III (11%), Stage II (5%), and Stage VI (5%), respectively. Baseline characters of Group I and Group II were similar [Table 1]. Group I showed a statistically significant increase in hair count, hair density, and anagen to telogen ratio (141 ±24, 215.14 ±34.84, and 8.09 ±3.47) as compared to Group II (113.17 ±21.84, 170.23 ± 32.95, and 5.33 ±3.62) at 3 months [Figure 1], [Figure 2] and [Table 1]. The improvement in terminal/vellus hair ratio was better in Group I as compared to Group II, but change was statistically insignificant [Table 1]. Hair diameter remained fairly constant in both groups over the follow-up of 3 months. On self-assessment questionnaire, the most common improved parameter was a decrease in hair fall (80% of patients), followed by improvement of hair texture (60% of patients), followed by change in volume (50% of patients), followed by growth of new hairs (40% of patients), and pigmentary changes (10% of patients). Global photography revealed mild-to-moderate improvement in hair profile in 77% of cases in Group I as compared to 30% in Group II at 3 months [Figure 3] and [Figure 4].

Figure 1: Comparative hair count of Group I (aPRP intervention group) and Group II (placebo group) at baseline, 1 month, and 3 months

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Figure 2: Comparative anagen/telogen ratio of Group I (aPRP intervention group) and Group II (Placebo group) at baseline, 1 month, and 3 months

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Table 1: Comparison of various hair parameters in Group I and Group II at baseline, 1, and 3 months

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Figure 3: Patient 11, left – pretreatment, right – 3-month posttreatment. 3-month follow-pictures showing noticeable improvement in hair profile in posterior half (Group I) as compared to anterior half (Group II)

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Figure 4: Patient 25, left – pretreatment, right – 3-month posttreatment. 3-month follow-up pictures showing noticeable improvement of hair profile in anterior half (Group I) as compared to posterior half (Group II)

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Discussion

PRP is arguably one of the most important biological agents in research today. Being autologous, rich in growth factors, and minimally invasive, its application has significantly increased in wound healing, various musculoskeletal pathologies, and hair restoration. It secretes different types of proteins (growth factors) on activation. These growth factors help in cell proliferation, chemotaxis, migration, cellular differentiation, and extracellular matrix synthesis. In 2006, Uebel et al. hypothesized that these growth factors act on dermal papilla, leading to an intense vascular endoneogenesis and the progression of new hairs to the anagen phase.^[5] Schiavone et al. revealed an average increase of hair shaft diameter after 6 months of initiation of PRP therapy.^[7] These studies provided a new perspective concerning the application of PRP for alopecia in both transplanted and nontransplanted patients.

Till date, dilemma still persist, how to use treatment in terms of number of sessions, time interval between them and at what dosage. In our study, we gave five cycles of PRP at interval of 2 weeks, whereas in previous studies, number of cycles varied from 1 to 6 weeks while time interval between consecutive cycles also varied from 2 week to 1.5 months.^{[7],[8],[9],[11]} Since the half-life of platelets is 7–10 days, and we need to maintain long-lasting stimulation of growth factors on hair follicles, so this 2-week interval between subsequent sessions appeared most appropriate. Previous studies had documented evidence of new hair growth as early as 1 week after first application of PRP and the effect peaked at about 1–3 months, so five cycles of PRP seemed appropriate. On the other hand, at present, there are many types of kits commercialized for the use of PRP, but there is a difference in the centrifugation guidelines that make comparison or evaluation of each study difficult. Variations in kits along with their high cost also make it difficult to choose one among many. In our study, we used manual two-step centrifugation for aPRP production. The efficacy of this protocol has been established in review analysis conducted by Dhurat and Sukesh.^[10] Such manual method of preparation is quite simple, economical, and cost-effective in developing countries like India.

Alves and Grimalt^[11] and Mapar et al.^[12] conducted half-headed trial, in which PRP was given on either right or left half of scalp, while placebo in other half. In our setup, it is difficult to convince patients for drug injection in one half and placebo in other. Hence, we designed a study, in which alopecia scalp was divided into anterior and posterior half for PRP and placebo injection. A similar design of the study had also been conducted by Cervelli et al.^[13] and Gentile et al.,^[14] but sample size of all these studies was quite small as compared to our study. Lack of randomization and blinding of treated and controlled groups were the shortcoming of our study as compared to above mentioned studies.

To use the mitogenic potential of growth factors for angiogenesis, fibroblast differentiation/proliferation, and collagen synthesis, platelet alpha-granules must be released by platelet activation. Borhan et al.^[15] used non-aPRP for treatment of AGA, considering that the centrifugation, intradermal injections (micro blood vessel injuries), and the contact with dermal fibroblasts would be sufficient to activate platelets and to release growth factors. Their study reported a mean increase of hair density by 2.9%. Cervelli et al.^[13] found 19% gain in mean hair density when they used aPRP in AGA patients, while Trink et al.^[16] noticed better results with aPRP in alopecia areata patients. In our study, we used aPRP as we wanted to ensure most optimum concentration of growth factors to hair follicles which in turn would show more efficacy. However, Gentile et al. investigated the effect of activated and non-aPRP on hair growth and found no difference in end results in both the groups.^[14] The results of our study revealed that the administration of aPRP led to a statistically significant increase in the mean hair density (26%), hair count (24%), and anagen/telogen ratio (47%) after 3 months when compared with baseline [Figure 5]. Regarding the mean total hair density, hair count, and anagen/telogen ratio, the increase on aPRP-treated side was also found to be statistically significant compared with that on the control side after 3 months. Circulating androgens, particularly DHT, bind to ARs in the dermal papilla and suppress pro-growth Wnt, Shh, and STAT3 signaling while activating inhibitory cascades.^[17] The growth factors found in PRP, including platelet-derived growth factor, transforming growth factor-beta, epidermal growth factor (EGF), insulin-like growth factor-1, and vascular endothelial growth factor, reinstate the necessary signaling pathways and gene transcription that result in cell survival, proliferation, and differentiation. They stimulate telogen-to-anagen transition, prolong anagen, and promote neoangiogenesis. In our study, placebo group also showed slight improvement, though not statistically significant, of all parameters. This might be due to diffusion of PRP into control areas as both areas were adjacent to each other. The results were similar to the study conducted by Alves and Grimalt.^[11]

Figure 5: Patient 20 (phototrichogram), left – pretreatment, right – 3-month posttreatment (Group I) showing improvement in diameter and density of hair

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In our study, global photography showed mild-to-moderate improvement in hair profile of aPRP-treated area in 73% of cases which was statistically significant as compared to 14% of cases in control area. Similarly, a study conducted by Anitua et al.^[18] also demonstrated improvement of hair profile on global photography at 1 year. Study conducted by Borhan et al.^[15] showed disappointing cosmetic results with global photography in spite of increased hair density. However, the overall satisfaction rate was good in patients with Grade II–III of alopecia, but patients with a higher grade of alopecia (Grade IV–VI) were not satisfied with this treatment as none of the patients showed marked improvement in hair profile. Most of these patients were looking for some other modalities of hair restoration in spite of getting full-session aPRP therapy.

This study has the limitation of having a follow-up of only 3 months which is too short period to draw final conclusions about the benefits and relapse after discontinuation of therapy. Furthermore, it would be a better study design, if we would have compared the effect of aPRP to some other medication or non-aPRP.

In our study, the same patient played the role of both treated and controlled groups. This possibly could have corrected the gender bias and bias of hair loss grading. Besides local and transitory pain during administration of aPRP and saline solution and postprocedural heaviness, no other major side effects were noted or reported during the 3-month follow-up.

Conclusion

This study suggests aPRP as a new treatment modality for AGA in early stages. Being autologous, it is safe and efficient alternative to other medical treatments. The use of aPRP does not appear to be associated with increased patient satisfaction in advanced cases of AA. It seems to have a stabilizing effect on hair fall and may be considered as measure to prevent the progression of alopecia in early stages. However, further controlled and randomized studies are needed to validate our findings in a larger cohort of patients with longer follow-up.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

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