ArticlePDF Available

Effects of carbon dioxide therapy on the healing of acute skin wounds induced on the back of rats

Authors:

Abstract and Figures

Purpose: To evaluate the healing effect of carbon dioxide therapy on skin wounds induced on the back of rats. Methods: Sixteen rats underwent excision of a round dermal-epidermal dorsal skin flap of 2.5 cm in diameter. The animals were divided into two groups, as follows: carbon dioxide group - subcutaneous injections of carbon dioxide on the day of operation and at three, six and nine days postoperatively; control group - no postoperative wound treatment. Wounds were photographed on the day of operation and at six and 14 days postoperatively for analysis of wound area and major diameter. All animals were euthanized on day 14 after surgery. The dorsal skin and the underlying muscle layer containing the wound were resected for histopathological analysis. Results: There was no statistically significant difference between groups in the percentage of wound closure, in histopathological findings, or in the reduction of wound area and major diameter at 14 days postoperatively. Conclusion: Under the experimental conditions in which this study was conducted, carbon dioxide therapy had no effects on the healing of acute skin wounds in rats.
Content may be subject to copyright.
334 - Acta Cirúrgica Brasileira - Vol. 28 (5) 2013
3 – ORIGINAL ARTICLE
WOUND HEALING
Effects of carbon dioxide therapy on the healing of acute
skin wounds induced on the back of rats1
Maria Vitória Carmo PenhavelI, Victor Henrique Tavares NascimentoII, Eliana Ferreira Ribeiro DurãesIII, Fabiana Pirani
CarneiroIV
, João Batista de SousaV
IGraduate student, School of Medicine, University of Brasilia(UnB), Brasilia-DF, Brazil. Acquisition and interpretation of data, technical procedures,
statistical analysis, manuscript writing.
IIGraduate student, School of Medicine, UnB, Brasilia-DF, Brazil. Acquisition and interpretation of data, technical procedures, statistical analysis,
critical revision.
IIIFellow Master degree, Postgraduate Program in Medical Sciences, School of Medicine, UnB, Brasilia-DF, Brazil. Technical procedures, statistical
analysis, interpretation of data, critical revision.
IVPhD, Associate Professor of Pathology, School of Medicine, UnB, Brasilia-DF, Brazil. Macroscopic and histopathological examinations, interpretation
of data, critical revision.
VPhD, Associate Professor of Surgery, School of Medicine, UnB, Brasilia-DF, Brazil. Intellectual and scientic content of the study, design the
protocol, technical procedures, interpretation of data, manuscript writing, critical revision.
ABSTRACT
PURPOSE: To evaluate the healing effect of carbon dioxide therapy on skin wounds induced on the back of rats.
METHODS: Sixteen rats underwent excision of a round dermal-epidermal dorsal skin ap of 2.5 cm in diameter. The animals were
divided into two groups, as follows: carbon dioxide group – subcutaneous injections of carbon dioxide on the day of operation and at
three, six and nine days postoperatively; control group – no postoperative wound treatment. Wounds were photographed on the day of
operation and at six and 14 days postoperatively for analysis of wound area and major diameter. All animals were euthanized on day
14 after surgery. The dorsal skin and the underlying muscle layer containing the wound were resected for histopathological analysis.
RESULTS: There was no statistically signicant difference between groups in the percentage of wound closure, in histopathological
ndings, or in the reduction of wound area and major diameter at 14 days postoperatively.
CONCLUSION: Under the experimental conditions in which this study was conducted, carbon dioxide therapy had no effects on the
healing of acute skin wounds in rats.
Key words: Wound Healing. Carbon Dioxide. Skin. Rats.
Effects of carbon dioxide therapy on the healing of acute
skin wounds induced on the back of rats
Acta Cirúrgica Brasileira - Vol. 28 (5) 2013 - 335
Introduction
Wound healing consists in a coordinated cascade of
cellular and molecular events that interact to allow tissue repair.
Tissue injury triggers healing and initiates a series of temporally
overlapping phases, including inammation, new tissue formation,
and tissue remodeling. Immediately after injury, the repair
process is initiated by the action of a number of growth factors,
cytokines, and products released from platelets and injured blood
vessels. After the formation of the blood clot, inammatory cells
invade the wound tissue and exert functions of protection against
contaminating microorganisms, being also an important source
of growth factors and cytokines, which initiate the proliferative
phase of wound healing. The proliferative phase starts with the
migration and proliferation of keratinocytes at the wound edge
and is followed by multiplication of dermal broblasts in the
neighborhood of the injured tissue. Subsequently, broblasts
begin to produce large amounts of extracellular matrix. Still in
the proliferative phase, the granulation tissue is formed, name
given by the granular appearance due to the presence of newly
formed capillaries, which are essential for the repair process.
Finally, a transition from granulation tissue to mature scar occurs,
characterized by continued collagen synthesis and degradation.
The scar tissue is mechanically insufcient and lacks epidermal
appendages1,2.
Since ancient times, man has tried to interfere in the tissue
repair process. In recent decades, much effort has been devoted
to identify substances and techniques in wound management
which are able to promote healing. The search for substances
with angiogenic activity has also been enhanced because of their
great potential for clinical application. However, due to the high
prevalence of complicated chronic ulcers, associated with organic
diseases and disorders, wound healing remains a challenging
clinical problem3.
The controlled injection of carbon dioxide (CO2) or CO2
therapy has been used as a simple, safe, biocompatible, low-cost
treatment option for wound management. CO2 therapy requires a
portable device that enables subcutaneous application of CO2 for
therapeutic purposes4.
The percutaneous route was the rst to be used by means
of CO2-enriched water baths to treat peripheral vascular diseases5.
Injections of CO2 in the skin and adipose tissue have been widely
used in aesthetic medicine with good results6-8. Studies have
demonstrated that CO2 plays a role in the recovery of granulation
tissue in chronic venous insufciency ulcers9, and stimulates
granulation in acute and chronic ulcers10.
The search for better clarication about the healing
process and the factors able to modify it is of fundamental
importance for daily surgical practice. Therefore researchers
remain extremely committed to the study of healing in different
experimental models11-13.
This study aimed to evaluate the healing effect of CO2
therapy on skin wounds induced in rats.
Methods
All experiments and procedures were carried out in
accordance with the ethical principals involving animals in research
dened by the Brazilian College for Animal Experimentation
(COBEA), afliated with the International Council for Laboratory
Animal Science, and conformed to the Brazilian guidelines
involving animals in research (Federal Law No. 6638 of 1979).
The study was approved by the Institutional Animal Care and Use
Committee of University of Brasilia, School of Medicine (protocol
no. 52439/2011).
Sixteen adult male Wistar rats (Rattus norvegicus
albinus, Rodentia mammalia), with initial body weight of 188 to
386g and about 60 days of age, were used in the study. For seven
days preoperatively, the animals were housed individually in home
cages, maintained on a 12:12 light/dark cycle at room temperature
and humidity, and had free access to water and specic chow
(Purina®, Labina Ltda., Campinas-SP, Brazil).
Experimental design
The animals were randomly divided into two groups of
eight individuals each to receive either subcutaneous injections
of CO2 on the day of operation and at three, six and nine days
postoperatively (CO2 group) or no postoperative wound treatment
(control group). All animals were killed on day 14 after surgery.
Surgical procedure
The animals were anesthetized with xylazine
hydrochloride (10 mg/kg body weight) and ketamine hydrochloride
(75 mg/kg body weight) intramuscularly. All surgical procedures
were performed by same team.
After an adequate level of anesthesia was obtained, the
animals were xed in the prone position on a surgical board. Before
surgical intervention, an area on the dorsal region of each rat was
shaved and aseptically prepared with 2% chlorhexidine gluconate.
All animals in both groups underwent the same surgical procedure.
The center of the shaved area was previously demarcated with
a dermatological metallic punch of 2.5 cm in diameter, and the
Penhavel MVC et al.
336 - Acta Cirúrgica Brasileira - Vol. 28 (5) 2013
excision of the skin ap was completed using a scalpel (Figure 1).
Hemostasis was achieved by digital compression of the bleeding
point with gauze. Then, animals in the experimental group received
CO2 injected subcutaneously into the wound edge by inserting the
needle directed toward the center of the wound (Figure 2). Animals
in the control group received no postoperative wound treatment.
FIGURE 1 - Skin excision. Detail of the “punched-out” area, which was
resected respecting the muscle layer.
FIGURE 2 - Carbon dioxide (CO2) therapy. Subcutaneous injection of
CO2 into the wound edge, with the needle directed toward the center of
the wound.
CO2 therapy was performed again at three, six and nine
days postoperatively in the experimental animals. All animals,
including control rats, were previously anesthetized on the days
mentioned above. Neither group received occlusive dressing
after application of treatment. At the end of the procedures, the
animals were placed back in their respective cages under the same
preoperative conditions.
Postsurgical wound progression
With the animals xed on the surgical board, minor
and major wound diameters were measured using calipers for
comparison with the standard initial measurement. At this moment,
wounds were photographed using a digital camera. This procedure
was performed on the day of operation and at six and 14 days
postoperatively. The images were transferred to ImageJ® software
and, after outlining the wound boundary using the polyline method
(demarcation of all points of the wound), all images were analyzed
for wound area and major diameter.
Collection of material for analysis
On the 14th postoperative day, all 16 animals were
anesthetized with ketamine and xylazine intramuscularly. Then,
a dorsal block containing the wound and the underlying muscle
layer was resected. The animals were killed with a lethal dose of
thiopental intraperitoneally (25 mg/kg). The specimens were xed
in formalin for histopathological analysis.
Histopathology
For histopathological analysis, parafn-embedded
specimens were stained with hematoxylin and eosin and examined
under an optical microscope. The amount of collagen, broblasts,
and polymorphonuclear and mononuclear inltration was
analyzed and graded on a scale of 0 to 3, with 0 indicating absence,
1 mild amount, 2 moderate amount, and 3 marked amount of the
analyzed parameter. Newly formed vessels were quantied in ve
high-power elds. The presence or absence of re-epithelialization,
foreign body, abscesses or hair follicles in the scar tissue were also
documented.
Statistical analysis
Data were analyzed using SigmaStat® version 3.5.
Wound areas and major diameters in experimental and control
groups, on each experimental day, were compared using one-way
analysis of variance (ANOVA). Fisher’s exact test and the chi-
square test were used to evaluate histological variables. The level
of signicance was set at p<0.05.
Results
Wound measurements
A comparison of wound areas between CO2 and control
groups, on day 0 and postoperative day 6, showed no statistically
signicant difference (p=0.220 and p=0.090, respectively),
although on postoperative day 6 the CO2 group showed a mean
wound area smaller than that of controls. On postoperative day
14, there was also no signicant difference in wound area between
groups (p=0.157) (Table 1).
Effects of carbon dioxide therapy on the healing of acute
skin wounds induced on the back of rats
Acta Cirúrgica Brasileira - Vol. 28 (5) 2013 - 337
TABLE 1 - Wound area (cm2) in control and carbon
dioxide (CO2) groups.
Experimental
day
Control (n=8)
Mean ± SD
CO2 (n=8)
Mean ± SD p
04.17±0.88 3.73±0.45 0.200
61.69 ±0.52 1.31±0.29 0.090
14 0.04±0.04 0.08±0.06 0.157
SD = standard deviation.
Regarding the measurement of wound major diameter,
there was no statistically signi cant difference between groups on
any experimental day.
The percentage of wound closure from day 0 to
postoperative day 6 showed no statistically signi cant difference
in the comparison between groups (p=0.182). However, a higher
percentage of wound closure was observed in the CO2 group
compared to the control group (64.8% and 59.5%, respectively).
Microscopic evaluation
Tables 2, 3 and 4 show the histological comparison
between groups on postoperative day 14, revealing no statistically
signi cant difference in any factor analyzed. Although not
statistically signi cant, the number of neovessels in the CO2
group, as observed in high-power microscopic elds, was larger
than that of controls (Figure 3).
TABLE 2 - Histological comparison between control
and carbon dioxide (CO2) groups on the 14th postoperative day.
Control
n=8
CO2
n=8
Mean Max/min Mean Max/min
Collagen 2.0 2/2 2.0 2/2
Fibroblasts 3.0 3/3 3.0 3/3
Mononuclear 2.0 2/2 2.0 2/2
Polymorphonuclear 2.0 2/2 2.0 2/2
Epithelial hyperplasia 1.4 2/1 1.9 2/1
CO2 X control, p=0.105.
TABLE 3 - Histological comparison between control
and carbon dioxide (CO2) groups on the 14th postoperative day
(2).
Control
n=8
CO2
n=8
Present Absent Present Absent
Hair follicle 0 8 0 8
Abscess 0 8 0 8
Foreign body 8 0 8 0
Epithelialization 4435
TABLE 4 - Comparison between control and carbon
dioxide (CO2) groups – quanti cation of newly formed vessels on
the 14th postoperative day.
Control (n=8)
Mean ± SD
CO2 (n=8)
Mean ± SD p
Vessel /  eld 24.50±9.24 37.87±20.12 0.110
SD = standard deviation.
A B
FIGURE 3 - Photomicrograph of repair tissue of rat skin – 14 days
postoperatively. A. Carbon dioxide group; B. Control group (H&E, 40x).
Greater amount of neovessels in A (arrow).
Discussion
The present study investigated the possible effect of
controlled subcutaneous injection of CO2 as an experimental
procedure on wounds induced on the back of rats. According to
available literature, CO2 therapy is a biocompatible treatment,
which is easy to perform, has properties of neoangiogenesis and
tissue repair, and contributes to healing in animal models and
humans.
The increase in CO2 tissue concentration in the
microcirculation, caused by controlled injection of this gas, leads to
a reduction in pH and an immediate increase in oxygen availability
to tissues through the Bohr effect. As a nal consequence of the
Penhavel MVC et al.
338 - Acta Cirúrgica Brasileira - Vol. 28 (5) 2013
reduction in pH and the release of mediators, vasomotor effects
on the microcirculation, neoangiogenesis and brogenesis in
the extracellular matrix are observed4,9. Ito et al.5 using CO2
percutaneously, provided experimental support for the clinical use
of CO2-enriched water bathing in the treatment of skin circulatory
disorders and ulcers.
According to Piazzolla at al.4, in a preliminary experience
with carbon dioxide therapy in the treatment of pressure ulcers in
a bedridden elderly patient, this treatment is especially benecial
for elderly adults with pressure ulcers due to immobility because
they may have injections at home, avoiding hospitalization.
Abramo and Teixeira9 administered CO2 therapy by
subcutaneous injection in 10 patients with chronic venous ulcers
of the lower limbs, with a mean ulcer duration of 12.5 months. The
authors used a 72-hour interval between CO2 application sessions,
similar to what was proposed in our study. Although the authors
found no signicant reduction in wound dimensions, all wounds
showed increased numbers of macrophages and broblasts and
improved granulation tissue after treatment.
Those and other studies supporting the efcacy of CO2
therapy have used this treatment in patients with chronic ulcers,
especially ulcers due to venous insufciency. Such results are
best explained by the effect of CO2 therapy on clinical conditions
or diseases that involve, in their pathophysiology, tissue hypo-
oxygenation9. These conditions differ from those investigated
in this study, which analyzed acute ulcers induced in healthy
animals. This may explain the discordant results, since there was
no statistical signicance when comparing wound area reduction
and histopathological parameters between the group receiving CO2
therapy and untreated control animals. Furthermore, despite using
the same treatment, a discrepancy between the results obtained
in chronic ulcers, of any etiology, and those in acute ulcers is
expected, since the latter have a different microenvironment
and healing pattern. Differences at the level of proinammatory
cytokines, proteases, cell mitosis and growth factors between a
healing ulcer and difcult-to-heal ulcers emphasize the need for
specic therapies for each type of wound14.
In the present study, the animals receiving CO2 therapy
showed greater neovascularization than controls, although without
statistical signicance. In agreement with this observation, Wollina
et al.10, studying patients with chronic ulcers of different origin
and patients with acute surgical wounds undergoing CO2 therapy,
found clinical evidence of improvement in granulation, even in
acute ulcers. Other studies have also highlighted the neoangiogenic
properties of CO2 therapy, even proposing the involvement of
vascular endothelial growth factor (VEGF) and nitric oxide (NO)
formation in this mechanism15.
Nevertheless, further studies involving a larger sample
of animals are warranted to clarify the benecial effect of CO2
therapy on acute ulcers. Serial wound biopsies may also be used in
order to evaluate the effect of CO2 therapy on each developmental
phase of wound healing.
Conclusion
Carbon dioxide therapy had no effects on the healing of
acute skin wounds induced in rats.
References
1. Mendonça RJ, Coutinho-Netto J. Aspectos celulares da cicatrização.
An Bras Dermatol. 2009;84(3):257-62.
2. Werner S, Grose R. Regulation of wound healing by growth factors
and cytokines. Physiol Rev. 2003;83(3):835-70.
3. Velnar T, Bailey T, Smrkolj V. The wound healing process: an
overview of the cellular and molecular mechanisms. J Int Med Res.
2009;37(5):1528-42.
4. Piazzolla LP, Louzada LL, Scoralick FM, Martins, ME, de Sousa,
JB. Preliminary experience with carbon dioxide therapy in the
treatment of pressure ulcers in a bedridden elderly patient. J Am
Geriatr Soc. 2012;60(2):378–9.
5. Ito T, Moore JI, Koss MC. Topical application of CO2 increases skin
blood ow. J Invest Dermatol. 1989;93(2):259-62.
6. Brandi C, D’Aniello C, Grimaldi L, Caiazzo E, Stanghellini E.
Carbon dioxide therapy: effects on skin irregularity and its use as a
complement to liposuction. Aesthetic Plast Surg. 2004;28(4):222–5.
7. Brandi C, D’Aniello C, Grimaldi L, Bosi B, Dei I, Lattarulo P,
Alessandrini C. Carbon dioxide therapy in the treatment of localized
adiposities: clinical study and histopathological correlations.
Aesthetic Plast Surg. 2001;25(3):170-4.
8. Nach R, Zandifar H, Grupta R, Hamilton JS. Subcutaneous
carboxytherapy injection for aesthetic improvement of scars. Ear
Nose Throat J. 2010;89(2):64-6.
9. Abramo AC, Teixeira TT. Carboinsuação em úlceras crônicas de
membros inferiores. Rev Bras Cir Plast. 2011;26(2):205-10.
10. Wollina U, Heinig B, Uhlemann C. Transdermal CO2 application in
chronic wounds. Int J Low Extrem Wounds. 2004;3(2)103–6.
11. Oliveira MVM, Brandão AM, Morais PHA, Silva NG, Silva SM,
Carneiro FP, Sousa JB. Effects of bromopride on abdominal wall
healing with induced peritoneal sepsis after segmental colectomy
and colonic anastomosis in rats. Acta Cir Bras. 2011;26(6):433-7.
12. Melo VA, Anjos DCS, Albuquerque Júnior R, Melo DB, Carvalho
FUR. Effect of low level laser on sutured wound healing in rats.
Acta Cir Bras. 2011;26(2):129-34.
13. Silva NG, Brandão AM, Oliveira MVM, Morais PHA, Silva SM,
Carneiro FP, Sousa JB. Inuence of metoclopramide on abdominal
wall healing in rats subjected to colonic anastomosis in the presence
of peritoneal sepsis induced. Acta Cir Bras. 2011;26(Supl 2):92-9.
14. Trengove NJ, Bielefeldt-Ohmann H, Stacey MC. Mitogenic activity
and cytokine levels in non-healing and healing chronic leg ulcers.
Wound Repair Regen. 2000;8(1):13-25.
15. Irie H, Tatsumi T, Takamiya M, Zen K, Takahashi T, Azuma A,
Tateishi K, Nomura T, Hayashi H, Nakajima N, Okigaki M,
Matsubara H. Carbon dioxide−rich water bathing enhances collateral
Effects of carbon dioxide therapy on the healing of acute
skin wounds induced on the back of rats
Acta Cirúrgica Brasileira - Vol. 28 (5) 2013 - 339
blood ow in Ischemic hindlimb via mobilization of endothelial
progenitor cells and activation of NO-cGMP system. Circulation.
2005;111(12):1523-9.
Correspondence:
João Batista de Sousa
Universidade de Brasília
Campus Universitário Darcy Ribeiro-Faculdade de Medicina
Laboratório de Cirurgia Experimental
70910-900 Brasília – DF Brasil
Tel.: (55 61)9984-2845
sousajb@unb.br
Received: January 23, 2013
Review: March 21, 2013
Accepted: April 22, 2013
Conict of interest: none
Financial source: none
1Research performed at Experimental Surgery Laboratory, School of
Medicine, University of Brasilia (UnB), Brazil.
... Somez et al. [22] conducted a study using controlled CO2 insufflation, but did not report needle size or angle of insertion. Penhavel et al. [36] conducted a randomized study with Wistar rats, but [37] reported having inserted the whole needle into the animal at a 90° angle to inject the gas into the hypodermic tissue. However, during our pilot study, when the 13 × 5 needle was inserted at a 90° angle to the tissue, it punctured the skin, hypodermis, and underlying muscle, reaching the animal's insides. ...
... A number of different flow parameters and puncture times have been used in different animal studies: 10-20 ml/min20, [38]; 30-50 ml/min, [41]; and 80 ml/min for 10 s [39]. However, many studies fail to mention the parameters used, including an investigation conducted by Penhavel et al. [36]. ...
... In the control group (without gas insufflation), as expected, all layers beyond the local epidermis were damaged where the needle was inserted, which could be explained by inflammatory infiltration [36]. ...
Article
Full-text available
Background Despite the indications of carboxitherapy in situations of ischemia and necrosis, there is still no scientific evidence to support its clinical applicability. The purpose of this study was to investigate the effect of controlled carbon dioxide insufflation on the viability of dorsal ischemic random skin flaps in rats. Methods A randomized, single-blind, experimental study was conducted. Thirty-six Wistar-EPM rats were randomly distributed into three groups: group 1 (control using ischemic random skin flap procedure without gas insufflation); group 2 (flap procedure with controlled carbon dioxide insufflation); and group 3 (flap procedure with controlled inert gas insufflation). In groups 2 and 3, gas was insufflated intradermally in rats for 5 s (150 ml/min) using a 0.2-mm needle inserted at a 90° angle to the skin at two points (one located 2.5 cm from the cranial base of the flap and the other 7.5 cm from the midline of the flap). This procedure was repeated for 7 consecutive days. Macroscopic analysis (necrotic area) was performed using the paper-template method proposed by Sasaki and Pang. Microscopic analysis of the vascularization process was performed using hematoxylin and eosin staining. Results No statistically significant differences were found for the parameter vascularization, inflammatory infiltrate, and percentage of necrotic area. Conclusions Controlled carbon dioxide insufflation did not have a significant impact on the viability of ischemic random skin flaps. Level of evidence: Not ratable.
... Multiple factors influence this process, including blood supply, defect size, tension, mobility, susceptibility to infection, type, and condition of the underlying tissue. 1 Excessive, continuous, and chronic inflammation may affect healing and prevent morphofunctional normality, resulting instead in the formation of disoriented connective tissue. This abnormal architecture reduces mechanical strength of tissues and leads to scar formation. ...
... treatments include hyaluronic acid, growth factors and blood components, stem cells, and adipose-derived stromal vascular fraction as well as negative-pressure wound therapy, ozone therapy, electrical stimulation, ultrasound, and phototherapy. [1][2][3][4][5][6][7][8] The variety of tested therapies highlights the level of interest in improving the healing of cutaneous lesions because of their frequency and potential challenging consequences. ...
Article
Full-text available
Objective To determine the effect of the Klox fluorescence biomodulation system (Phovia) on the healing of surgical wounds. Study design Prospective, blinded, controlled clinical trial. Sample population Healthy dogs undergoing orthopedic surgery (n = 10). Methods Half of the length of each surgical wound was treated with Phovia, and the remaining 50% was treated with saline solution on the first day after surgery and every 3 days until day 13. Wound healing of treated and control areas within each wound was evaluated via macroscopic assessment and histological and immunohistochemical analysis of treated and control wounds. Results The areas treated with Phovia achieved lower histology scores (P = .001), consistent with complete re‐epithelialization, less inflammation of the dermal layer, and greater and more regular deposition of collagen. According to immunohistochemistry, expression of factor VIII, epidural growth factor, decorin, collagen III, and Ki67 was increased in treated compared with untreated tissues. Conclusion Phovia therapy improved re‐epithelialization, decreased dermal inflammation, and improved matrix formation in uncomplicated cutaneous incisional wounds by regulating the expression of key biological mediators. Clinical significance Phovia may be a beneficial adjunct to promote the healing of incisional wounds.
... Consequently, this points to CO 2 not only as a by-product of aerobic respiration but also to its role as a gasotransmitter in suppressing inflammatory responses. In the skin, CO 2 has been shown to improve wound healing and seasonal barrier dysfunction [23][24][25] ; inflammation is known to be strongly involved in skin wound healing and barrier functions as well as the circulatory system. The wound-healing process consists of three phases: the inflammatory phase, proliferative phase, and stable phase; this process may be delayed when an excessive inflammatory response is triggered during the inflammatory phase 26 . ...
Article
Full-text available
Carbon dioxide (CO2) is the predominant gas molecule emitted during aerobic respiration. Although CO2 can improve blood circulation in the skin via its vasodilatory effects, its effects on skin inflammation remain unclear. The present study aimed to examine the anti-inflammatory effects of CO2 in human keratinocytes and skin. Keratinocytes were cultured under 15% CO2, irradiated with ultraviolet B (UVB), and their inflammatory cytokine production was analyzed. Using multiphoton laser microscopy, the effect of CO2 on pH was observed by loading a three-dimensional (3D)-cultured epidermis with a high-CO2 concentration formulation. Finally, the effect of CO2 on UVB-induced erythema was confirmed. CO2 suppressed the UVB-induced production of tumor necrosis factor-α (TNFα) and interleukin-6 (IL-6) in keratinocytes and the 3D epidermis. Correcting medium acidification with NaOH inhibited the CO2-induced suppression of TNFα and IL-6 expression in keratinocytes. Moreover, the knockdown of H+-sensing G protein-coupled receptor 65 inhibited the CO2-induced suppression of inflammatory cytokine expression and NF-κB activation and reduced CO2-induced cyclic adenosine monophosphate production. Furthermore, the high-CO2 concentration formulation suppressed UVB-induced erythema in human skin. Hence, CO2 suppresses skin inflammation and can be employed as a potential therapeutic agent in restoring skin immune homeostasis.
... It also permits the release of growth factor for angiogenesis 18 . Some studies showed that vasodilation is mediated by nitric oxide, which induces angiogenesis and suppresses vasoconstriction in response to cold by keeping the patients' feet warm 19 . ...
Article
Full-text available
Background: Diabetic foot ulcer (DFU) considers one of the most anxious complications associated with uncontrolled diabetes mellitus. The aim of this study is to evaluate carbon dioxide therapy on patients with DFU and compare with patients on traditional therapy. Methods: 100 patients with DFU (mean age 51.6+ 5.65 years; 56men) divided into two groups ,traditional therapy group will be treated using advance dressings and antibiotics, while CO2 therapy group will be treated using advance dressings and CO2 therapy. ABI and Doppler measured Blood flow to the affected foot; also, the size, color and sensation of the ulcerative area were all evaluated and compared between two groups. Results: Results showed improvement of blood flow to the affected foot as well as improvement in the sensation , size and color of the ulcerative area .Conclusion: effectiveness of this method in the treatment of ischemic feet, and indicates it is potential utility as a form of physiotherapy in the treatment of ischemic feet .
... Once anesthetized, each animal was placed on the surgical board in prone position. After trichotomy, the surgical technique started, with the same standardization for the animals in all groups, as we previously described in another publication 17 . The center of the epilated region was previously marked with a metallic, 2.5 cm diameter dermatological punch, and the excision of the skin was completed with a scalpel (figure 1). ...
Article
Full-text available
Objective: to evaluate the effect of topical delivery of latex cream-gel in acute cutaneous wounds induced on the back of rats. Methods: we subjected sixteen rats to dermo-epidermal excision of a round dorsal skin flap, with 2.5cm diameter. We divided the animals into two groups: Latex Group: application of cream-gel-based latex throughout the wound bed on postoperative days zero, three, six and nine; Control group: no treatment on the wound. Photographs of the lesions were taken on the procedure day and on the 6th and 14th postoperative days, for analyzing the area and the larger diameter of the wound. We carried out euthanasia of all animals on the 14th postoperative day, when we resected he dorsal skin and the underlying muscle layer supporting the wound for histopathological study. Results: there was no statistically significant difference in the percentage of wound closure, in the histopathological findings or in the reduction of the area and of the largest diameter of the wounds among the groups studied on the 14th postoperative day. Conclusion: according to the experimental conditions in which the study was conducted, latex cream-gel did not interfere in the healing of acute cutaneous wounds in rats.
... Experiments on rat skin showed that subcutaneous injection of CO2 decreased the amount of Substance P and pro-Calcitonin Gene-Related Peptide (15 kDa) neuropeptides in rat skin, with no effects on the healing of acute skin wounds. 6,7 The aim of the study was to compare the tolerability, cosmetic effect, and correlations of hyaluronic filler and carbon dioxide injections in the cosmetic correction of nasolabial folds. ...
Article
Background: The main application of hyaluronic acid filling, in esthetic medicine, is the augmentation of soft tissues. The carbon dioxide therapy, instead, improves quality and elasticity of the dermis and increases the oxygen release to the tissue through an enhancing of the Bohr's effect. The aim of the study was to compare the efficacy, tolerability, and effect duration of hyaluronic acid fillers and the use of carbon dioxide therapy plus hyaluronic acid in the cosmetic correction of nasolabial folds. Materials and methods: Forty healthy female patients received a blinded and randomized treatment on nasolabial folds (hyaluronic acid in group A and hyaluronic acid plus subcutaneous injections of carbon dioxide in group B) for cosmetic correction of the nasolabial folds. The results were evaluated by two blinded plastic surgeons after the implant (1 week, 4 and 6 months) using a 1-5 graduated scale (GAIS), and at the same time, each patient was asked to express her opinion about the cosmetic result. Results: Any long-term adverse reaction was reported. The blinded evaluation at 4 and 6 months from the implant shows in all patients a maintenance of a good cosmetic result higher for the side treated with carbon dioxide therapy plus hyaluronic acid. Conclusions: At the control visit, 6 months after the treatment, the patients treated with hyaluronic acid plus carbon dioxide therapy maintain a satisfactory esthetic result while the nasolabial fold treated only with hyaluronic acid shows, in almost all patients, a come back to pretreatment appearance.
... The controlled injection of carbon dioxide (CO 2 ) therapy has been used as a simple, safe, biocompatible, low-cost treatment option for wound management. CO 2 therapy requires a portable device that enables subcutaneous application of CO 2 for therapeutic purposes 1 . Carbon dioxide administered percutaneously promotes dilation of precapillary arterioles, reperfusion of occluded capillaries, reduction in blood viscosi- ...
Article
Full-text available
OBJECTIVE: The purposes of this study was to assess the effect of repeated subcutaneous injections of CO2 on adipose tissue graft survival in immunosuppressed female nude mice. The authors designed an experimental study using volume measures, histopathological analysis and nuclear magnetic resonance of fat graft. The effect of repeated subcutaneous injection of CO2 is not yet investigated MATERIALS AND METHODS: Approximately 0.5 ml of human fat were transplanted in a group of female nude mice. The mice were treated with 3 injections of 80 µl each carbon dioxide (total 240 µl) for 7 weeks. Initially, in vivo measurements were conducted and subsequently a comprehensive histopathological analysis was performed. RESULTS: The presence of inflammation was graded absent to minimal in animals treated with CO2 while a minimal to moderate grade was assigned to the control group. CONCLUSIONS: CO2 injection enhances the inflammatory response of the implanted tissue and reduces the reabsorption rate. The treatment may improve the graft survival in a more prolonged time-frame.
... Some studies showed that vasodilation is mediated by nitric oxide which induces angiogenesis and suppresses vasoconstriction in response to cold by keeping the patients' feet warm [26]. Table 1 and Figure 1 showed improvement in the blood flow expressed by mean Doppler pressure pre and post treatment significantly (p > 0.5). ...
Article
Full-text available
Ulceration of diabetic foot represents one of the most concerning complications associated with uncontrolled blood sugar in diabetes mellitus. The aim of this study is to evaluate the condition of twenty-two diabetic patients with different degrees of ulceration in their feet after daily secessions of carbon dioxide therapy. Blood flow to the affected foot was measured by Doppler; also the size, color, degree of ulceration and sensation of the ulcerative area were all evaluated. Results showed improvement of blood flow to the affected foot as well as improvement in the sensation and color of the ulcerative area. It was concluded that carbon dioxide therapy of diabetic foot was promising and needed thorough investigation to be brought widely into application.
Article
Full-text available
O processo cicatricial compreende uma sequência de eventos moleculares e celulares que interagem para que ocorra a restauração do tecido lesado. Desde o extravasamento de plasma, com a coagulação e agregação plaquetária até a reepitelização e remodelagem do tecido lesado o organismo age tentando restaurar a funcionalidade tecidual. Assim, este trabalho abrange os diversos aspectos celulares envolvidos no processo cicatricial, bem como os principais medicamentos utilizados no tratamento de patologias relacionadas às deficiências na cicatrização. São abordados também, os aspectos econômicos referentes, sobretudo, às feridas crônicas de pés diabéticos.Wound healing is a dynamic interactive process that involves a sequence of molecular and cellular events. Recent advances in cellular and molecular biology have greatly expanded our understanding of the biological process involved in wound repair and tissue regeneration. From plasma extravasation, with coagulation and platelet aggregation, to reepithelialization and remodeling of injured tissue, the organism acts by trying to restore functionality tissue. Thus, the present study encompasses several cellular aspects involved in the wound healing process, as well as the main drugs used in treating the pathology related to wound healing complications. Economic aspects are also addressed, mainly related to chronic wounds of diabetic feet.
Article
Full-text available
The authors report their experience using carbon dioxide (CO2) therapy for the treatment of 48 female patients presenting adipose accumulations, located on the thighs, knees, and/or abdomen; a Carbomed Programmable Automatic Carbon Dioxide Therapy apparatus was used. In light of the effects of CO2 on the microcirculation recently described in the literature, we expected this gas, which we administered subcutaneously, to positively affect the physiological oxidative lipolytic process. The aim of our study was to evaluate the effect of this therapy on localized adiposities. As such, we describe the method we used and report the results observed in the areas treated (in terms of reduction in maximum circumference) as well as side effects. Furthermore, we assessed the effect of subcutaneous administration of CO2 on the microcirculation by showing changes in the Laser Doppler signal and in the concentration of transcutaneous oxygen tension (tcPO2). Pre- and posttreatment biopsies of tissues were performed in seven patients in order to study the changes induced by the use of CO2 on both adipose and connective tissues. All data obtained were statistically analyzed; values of P < 0.05 were considered significant.
Article
Full-text available
To evaluate the effects of metoclopramide on abdominal wall healing in rats in the presence of sepsis. 40 rats divided into two groups of twenty animals, subdivided into two subgroups of 10 animals each: group (E) - treated with metoclopramide, and saline-treated control group. The two groups were divided into subgroups of 10 to be killed on the 3rd day (n = 10) or day 7 (n = 10) after surgery. Sepsis was induced by cecal ligation and puncture. We performed also the section and anastomosis in left colon. The synthesis of the abdominal wall was made with 3-0 silk thread. We measured the breaking strength of the abdominal wall and made the histopathological evaluation. on 3rd day postoperative, the average breaking strength in the E group was 0.83 ± 0.66 and in group C was 0.35 ± 0.46 (p = 0.010). On the seventh day, the breaking strength in group E was 11.44 ± 5.07, in group C 11.66 ± 7.38 (p = 1.000). The E7 group showed lower inflammatory infiltration, foreign body reaction, fibrin than control. animals treated with metoclopramide had a higher resistance of the abdominal wall on the 3rd postoperative day.
Article
Full-text available
Evaluate the effects of bromopride on abdominal wall healing of rats with induced peritoneal sepsis after segmental colectomy and colonic anastomosis. Forty rats underwent sectioning of the left colon and end-to-end anastomosis and were divided into two groups of 20 animals for the administration of bromopride (bromopride group - B) or saline solution (control group - C). Each group was divided into subgroups of 10 animals each to be killed on the third (GB3 and GC3) or seventh postoperative day (GB7 and GC7). It was analyzed the following characteristics: breaking strength of the abdominal wall's wound; surgical and histopathological features of the abdominal wall; and clinical features of the rats. There was no difference between the groups in relation to the weight of the rats and the breaking strength of the abdominal wall's wound. The GB7 group presented less edema and less quantity of fibrin during histopathological evaluation compared to the GC7 group. Bromopride did not have harmful effects on the healing of abdominal wall in rats.
Article
Full-text available
To evaluate the effect of low-level laser therapy (LLLT) ë904 nm on healing of surgical wounds in rats. Forty male Wistar rats were used, divided into four groups, underwent incision along the lines Alba covering skin, subcutaneous and muscle abdominal, sutured continuously for nylon 5-0. Eight and fifteen days after the surgery process, the repairing area was removed and histological sections were stained with hematoxylin-eosin to assess cellularity inflammatory, Masson's Trichrome and Picrossirus to quantify the collagen fibers and immunohistochemical technique for counting newly formed vessels. The data were compared statistically using analysis of variance ANOVA, with a "post-hoc Tukey test, p <0.05. Low-level laser therapy reduced the intensity of the inflammatory reaction and influenced the dynamic of the immunoinflammatory response by inducing switching of the leukocyte infiltration pattern (neutrophilic to lymphoplasmacytic infiltration). Also stimulate the deposition and enhance the organization of collagen fibers, featuring a delicate collagen type III. Furthermore, it appeared to a significant increase in the average number of newly formed vessels (p = 0.00 and p = 0.02, respectively). Low-level laser therapy resulted in modulate of the inflammatory response, enhanced deposition of collagen fibers and increase in the average number of newly formed vessels.
Article
Werner, Sabine, and Richard Grose. Regulation of Wound Healing by Growth Factors and Cytokines. Physiol Rev 83: 835–870, 2003; 10.1152/physrev.00032.2002.—Cutaneous wound healing is a complex process involving blood clotting, inflammation, new tissue formation, and finally tissue remodeling. It is well described at the histological level, but the genes that regulate skin repair have only partially been identified. Many experimental and clinical studies have demonstrated varied, but in most cases beneficial, effects of exogenous growth factors on the healing process. However, the roles played by endogenous growth factors have remained largely unclear. Initial approaches at addressing this question focused on the expression analysis of various growth factors, cytokines, and their receptors in different wound models, with first functional data being obtained by applying neutralizing antibodies to wounds. During the past few years, the availability of genetically modified mice has allowed elucidation of the function of various genes in the healing process, and these studies have shed light onto the role of growth factors, cytokines, and their downstream effectors in wound repair. This review summarizes the results of expression studies that have been performed in rodents, pigs, and humans to localize growth factors and their receptors in skin wounds. Most importantly, we also report on genetic studies addressing the functions of endogenous growth factors in the wound repair process.
Article
The cause of impaired healing in chronic leg ulcers is not known. However, recent attempts to modify the healing process have focused on adding growth factors to stimulate healing and have failed to produce dramatic improvements in healing. This study used a unique model of chronic wound healing in humans to obtain wound fluid samples from chronic venous leg ulcers that had changed from a nonhealing to a healing phase. These samples were used to assess cytokine and growth factor levels, and mitogenic activity in these nonhealing and healing chronic wounds. The pro-inflammatory cytokines interleukin-1, interleukin-6 and tumor necrosis factor-αwere found to be present in significantly higher concentrations in wound fluid from nonhealing compared to healing leg ulcers. There were detectable levels but, no significant change in the levels of platelet derived growth factor, epidermal growth factor, basic fibroblast growth factor or transforming growth factor-βas ulcers healed. Wound fluid was added to fibroblasts in vitro to assess mitogenic activity. There was a significantly greater proliferative response to healing wound fluid samples compared to nonhealing samples. These results suggest that healing may be impaired by inflammatory mediators rather than inhibited by a deficiency of growth factors in these chronic wounds.
Article
This is a case report of a 60 year old female who had a previous rhytidectomy scar. She was treated with a process called carboxytherapy that resulted in great improvement of the scars with minimal to no discomfort or down time. Copyright © 2010 The American Laryngological, Rhinological, and Otological Society, Inc.