Ingesting carbonated water post-exercise in the heat transiently ameliorates hypotension and enhances mood state.

The objective was to assess if post-exercise ingestion of carbonated water in a hot environment ameliorates hypotension, enhances cerebral blood flow and heat loss responses, and positively modulates perceptions and mood states. Twelve healthy, habitually active young adults (five women) performed 60 min of cycling at 45% peak oxygen uptake in a hot climate (35°C). Subsequently, participants consumed 4°C carbonated or non-carbonated (control) water (150 and 100 mL for males and females regardless of drink type) at 20 and 40 min into post-exercise periods. Mean arterial pressure decreased post-exercise at 20 min only (P = 0.032) compared to the pre-exercise baseline. Both beverages transiently (∼1 min) increased mean arterial pressure and middle cerebral artery mean blood velocity (cerebral blood flow index) regardless of post-exercise periods (all P ≤ 0.015). Notably, carbonated water ingestion led to greater increases in mean arterial pressure (2.3 ± 2.8 mmHg vs. 6.6 ± 4.4 mmHg, P < 0.001) and middle cerebral artery mean blood velocity (1.6 ± 2.5 cm/s vs. 3.8 ± 4.1 cm/s, P = 0.046) at 20 min post-exercise period compared to non-carbonated water ingestion. Both beverages increased mouth exhilaration and reduced sleepiness regardless of post-exercise periods, but these responses were more pronounced with carbonated water ingestion at 40 min post-exercise (mouth exhilaration: 3.1 ± 1.4 vs. 4.7 ± 1.7, P = 0.001; sleepiness: -0.7 ± 0.91 vs. -1.9 ± 1.6, P = 0.014). Heat loss responses and other perceptions were similar between the two conditions throughout (all P ≥ 0.054). We show that carbonated water ingestion temporarily ameliorates hypotension and increases the cerebral blood flow index during the early post-exercise phase in a hot environment, whereas it enhances mouth exhilaration and reduces sleepiness during the late post-exercise phase.

Stimulus-Response Plots as a Novel Bowel-Sound-Based Method for Evaluating Motor Response to Drinking in Healthy People.

Constipation is a common gastrointestinal disorder that impairs quality of life. Evaluating bowel motility via traditional methods, such as MRI and radiography, is expensive and inconvenient. Bowel sound (BS) analysis has been proposed as an alternative, with BS-time-domain acoustic features (BSTDAFs) being effective for evaluating bowel motility via several food and drink consumption tests. However, the effect of BSTDAFs before drink consumption on those after drink consumption is yet to be investigated. This study used BS-based stimulus-response plots (BSSRPs) to investigate this effect on 20 participants who underwent drinking tests. A strong negative correlation was observed between the number of BSs per minute before carbonated water consumption and the ratio of that before and after carbonated water consumption. However, a similar trend was not observed when the participants drank cold water. These findings suggest that when carbonated water is drunk, bowel motility before ingestion affects motor response to ingestion. This study provides a non-invasive BS-based approach for evaluating motor response to food and drink, offering a new research window for investigators in this field.

A preliminary investigation of the impact of artificially carbonated water bathing on canine skin physiological and barrier function.

BACKGROUND: The effect of carbon dioxide (CO2 )-rich water bathing on the skin has been studied extensively in humans. However, there have been few studies evaluating the impact of CO2 -rich water bathing on canine skin physiology and barrier functions. OBJECTIVES: To evaluate the impact of artificially carbonated water (ACW) bathing on skin parameters in healthy beagles. ANIMALS: Six healthy beagles with no history of skin disease. MATERIALS AND METHODS: Body temperature, skin temperature, transepidermal water loss (TEWL), skin hydration and skin blood flow were evaluated before and after single ACW bathing (37°C, 20 min) with a CO2 concentration of >1000 ppm. RESULTS: After ACW bathing, skin blood flow significantly increased (p < 0.0001), yet there were no significant changes in body temperature (p = 0.3124), skin temperature (p = 0.4911), TEWL (p = 0.5167) or skin hydration (p = 0.3084). There were no adverse events during the trials. CONCLUSIONS AND CLINICAL IMPORTANCE: Artificially carbonated water water bathing could potentially increase skin blood flow without affecting skin temperature, body temperature and skin barrier function in dogs, similar to its effects in humans.

Millennial drinks: acidity, fluoride content, and enamel softening.

This in vitro study aimed to evaluate the acidity and fluoride content of beverages commonly consumed by millennials and the enamel-softening effect of these drinks on tooth enamel. The study included 13 beverages in 4 categories: energy (sports) drink, flavored sparkling water, kombucha, and other (an unsweetened iced tea, a vegetable-fruit juice blend, and a soft drink). The acidity was measured with a pH/ion meter, and the fluoride concentration was measured with a combined fluoride electrode coupled to the meter (n = 10 measurements per beverage). The Vickers hardness number of extracted molars was measured before and after a 30-minute immersion in 4 representative beverages via 2 immersion protocols (n = 10 per beverage per protocol): (1) immersion in the beverage only and (2) immersion alternating between the beverage and artificial saliva every other minute. The pH and fluoride concentrations of the beverages ranged from 2.652 to 4.242 and from 0.0033 to 0.6045 ppm, respectively. One-way analysis of variance (ANOVA) revealed that all differences between beverages in pH values were statistically significant, as were the majority of differences in fluoride concentrations (P < 0.001). The beverages and the 2 immersion methods significantly affected enamel softening (2-way ANOVA, P = 0.0001 to 0.033). The representative energy drink (pH 2.990; 0.0102 ppm fluoride) caused the greatest enamel softening followed by the representative kombucha (pH 2.820; 0.2036 ppm fluoride). The representative flavored sparkling water (pH 4.066; 0.0098 ppm fluoride) caused significantly less enamel softening than the energy drink and kombucha. A root beer (pH 4.185; 0.6045 ppm fluoride) had the least enamel softening effect. All tested beverages were acidic and had a pH below 4.5; only some contained fluoride. Flavored sparkling water, likely due to its higher pH, caused less enamel softening than the tested energy drink and kombucha. The fluoride content of kombucha and root beer lower their enamel-softening effects. It is imperative that consumers be aware of the erosive potential of beverages they consume.

The pH of bottled water commercially available in Australia and its implications for oral health.

With a higher pH level and being unlikely to erode the tooth, bottled water has been considered a safe alternative to acidic beverages. However, recent studies have reported some bottled water products in different countries to be acidic. The present paper aimed to examine the pH values of 42 bottled waters commercially available in Australia, using a pH meter and probe, and classify their risks to cause erosive tooth wear in comparison with the critical pH of enamel and dentine. Of the 42 bottled water samples collected, 81.0 and 73.8% were considered erosive to tooth dentine and enamel, respectively. Flavoured waters were the most acidic, followed by sparkling waters, spring waters, artesian waters, mineral waters, and alkaline waters. All sparkling waters and flavoured waters showed an erosive risk to the enamel and dentine. A portion of spring waters and artesian waters was also acidic enough to erode tooth structures. The findings of this work were of concern given the risk of sustaining erosive tooth wear from consuming bottled waters. Health promotion strategies including public awareness and education on oral health consequences related to the consumption of bottled water are needed. Future epidemiological and in vivo investigations are also warranted.

Effects of Carbonated Water Concentration on Swallowing Function in Healthy Adults.

In this study, we investigated the effects of carbonated water concentration on swallowing function using surface electromyography (sEMG). Healthy subjects (n = 52, 26.77 ± 3.21 years old) were asked to perform two swallows each of noncarbonated water, low-concentration carbonated water, medium-concentration carbonated water, and high-concentration carbonated water. Onset time, the mean sEMG activity amplitude, and duration of muscle activity in each swallow were measured and analyzed for orbicularis oris, masseter, submental muscle complex and infrahyoid muscles. Onset time significantly decreased and mean sEMG activity amplitude significantly increased with carbonation concentration. Therefore, stimulation with carbonation can be effective for modulating a faster and stronger swallow in the oral and pharyngeal phases of swallowing, and its effect on amplitude was greater in the oral phase than in the pharyngeal phase.Clinical Trials Registration This study is registered with Clinical Research Information Service (KCT0005925).

Clinical efficacy of artificially carbonated water bathing on superficial bacterial folliculitis in dogs.

BACKGROUND: Bathing with artificially carbonated water is reported to be a valuable therapeutic option for various human skin disorders. OBJECTIVE: To evaluate the clinical efficacy of artificially carbonated water bathing on superficial bacterial folliculitis (SBF) caused by Staphylococcus pseudintermedius (SP) in dogs. ANIMALS: Nineteen dogs with SBF from whom SP was isolated from skin lesions were enrolled. METHODS AND MATERIALS: Dogs with SBF were allocated randomly to either the artificially carbonated water bathing group or the control group bathed with tap water. The dogs were bathed with the designated water type on day (D)0, D7 and D14. Clinical scores and skin surface pH were evaluated on D0 and D21. Colony forming unit (cfu) assays were performed in vitro to investigate whether the artificially carbonated water affected growth of clinical SP isolates. RESULTS: The mean rate of improvement in the clinical scores was significantly higher in the carbonated water group than in the control group. Dogs bathed with carbonated water exhibited significant decreases in their skin surface pH after bathing; dogs bathed with tap water did not. No dogs experienced significant adverse events. The cfus of SP incubated in vitro with artificially carbonated water did not significantly differ from those incubated with tap water. CONCLUSION: Bathing with artificially carbonated water might be an effective and safe adjunctive therapy for canine SP-induced SBF.

Bacillus subtilis: As an Efficient Bacterial Strain for the Reclamation of Water Loaded with Textile Azo Dye, Orange II.

The azo dye orange II is used extensively in the textile sector for coloring fabrics. High concentrations of it are released into aqueous environments through textile effluents. Therefore, its removal from textile wastewater and effluents is necessary. Herein, initially, we tested 11 bacterial strains for their capabilities in the degradation of orange II dye. It was revealed in the preliminary data that B. subtilis can more potently degrade the selected dye, which was thus used in the subsequent experiments. To achieve maximum decolorization, the experimental conditions were optimized whereby maximum degradation was achieved at: a 25 ppm dye concentration, pH 7, a temperature of 35 °C, a 1000 mg/L concentration of glucose, a 1000 mg/L urea concentration, a 666.66 mg/L NaCl concentration, an incubation period of 3 days, and with hydroquinone as a redox mediator at a concentration of 66.66 mg/L. The effects of the interaction of the operational factors were further confirmed using response surface methodology, which revealed that at optimum conditions of pH 6.45, a dye concentration of 17.07 mg/L, and an incubation time of 9.96 h at 45.38 °C, the maximum degradation of orange II can be obtained at a desirability coefficient of 1, estimated using the central composite design (CCD). To understand the underlying principles of degradation of the metabolites in the aliquot mixture at the optimized condition, the study steps were extracted and analyzed using GC-MS(Gas Chromatography Mass Spectrometry), FTIR(Fourier Transform Infrared Spectroscopy), 1H and carbon 13 NMR(Nuclear Magnetic Resonance Spectroscopy). The GC-MS pattern revealed that the original dye was degraded into o-xylene and naphthalene. Naphthalene was even obtained in a pure state through silica gel column isolation and confirmed using 1H and 13C NMR spectroscopic analysis. Phytotoxicity tests on Vigna radiata were also conducted and the results confirmed that the dye metabolites were less toxic than the parent dye. These results emphasize that B. subtilis should be used as a potential strain for the bioremediation of textile effluents containing orange II and other toxic azo dyes.

Study on the performance of sewage sludge biochar modified by nZVI to remove Cu(II) and Cr(VI) in water.

In this study, to simultaneously dispose of sludge and wastewater containing heavy metals, sludge biochar loaded with nano zero-valent-iron (nZVI) was prepared at 700 °C (nBC700) to remove Cr(VI) and Cu(II). The results showed the removal capacity of biochar was greatly improved by loading nZVI, and the adsorption capacities of biochar for Cu(II) and Cr(VI) increased by 251.96% and 205.18%. Pseudo-second-order kinetic and Sips isotherm models were fitted to the removal processes. Intraparticle diffusion models showed the removal process was controlled by surface diffusion and intraparticle diffusion. Competitive experiments showed Cr(VI) can compete with Cu(II) for active sites, but Cr(VI) was more easily removed by nBC700 through cation bridge. The removal mechanism illustrated removing Cu(II) mainly depended on complex precipitation, followed by reduction reaction, while Cr(VI) was on the contrary. This work provided effective data for sludge disposal and heavy metal removal.

Plasma metabolomics analysis of the effects of drinking soda water on hyperglycemia mice using ultrahigh-pressure liquid chromatography-quadrupole-time of flight-mass spectrometry.

The aim of this study was to evaluate the effects of a natural soda water [Shi Han Quan (SHQ)] on hyperglycemia and plasma metabolic profiling and explore the mechanism using metabolomics techniques. Kun-Ming mice weighing 26 ± 2 g were used for the hyperglycemia animal model with alloxan and divided into control, hyperglycemia (HG), and HG + SHQ soda water (SHQ) groups. The experiment lasted for 30 days. The plasma metabolomic profiling of mice was determined using ultrahigh-pressure liquid chromatography-quadrupole-time of flight-mass spectrometry. After the mice drank SHQ soda water, the levels of insulin and blood glucose were significantly lower in the SHQ group compared with the control group, and the level of insulin sensitivity [insulin sensitivity index (ISI)] was significantly higher in the SHQ group compared with the HG group. The mice in the different groups after SHQ intervention could be separated into distinct clusters, and nine major plasma metabolites with significant differences between groups were found closely associated with blood glucose and ISI. The metabolic pathway analysis of these metabolites involved abnormal fatty acid oxidation and phospholipid, acylcarnitine, and corticoid metabolism. The results suggested the metabolic changes and possible mechanism of SHQ improving the alloxan-induced HG, and the findings provided insights into the prevention and control of HG and diabetes.

Ethanol as a Probe for the Mechanism of Bubble Nucleation in the Diet Coke and Mentos Experiment.

The Diet Coke and Mentos experiment involves dropping Mentos candies into carbonated beverages to produce a fountain. This simple experiment has enjoyed popularity with science teachers and the general public. Studies of the physicochemical processes involved in the generation of the fountain have been largely informed by the physics of bubble nucleation. Herein, we probe the effect of ethanol addition on the Diet Coke and Mentos experiment to explore the impact that beverage surface tension and viscosity have on the heights of fountains achieved. Our results indicate that current descriptions of the effects of surface tension and viscosity are not completely understood. We also extend and apply a previously reported, simplified version of Brunauer-Emmett-Teller theory to investigate kinetic and mechanistic aspects of bubble nucleation on the surface of Mentos candies in carbonated beverages. A combination of this new theory and experiment allows for the estimation that the nucleation sites on the Mentos candy that catalyze degassing are 1-3 µm in size, and that between 50,000 and 300,000 of these sites actively nucleate bubbles on a single Mentos candy. While the methods employed are not highly sophisticated, they have potential to stimulate fresh investigations and insights into bubble nucleation in carbonated beverages.

Unveiling Carbon Dioxide and Ethanol Diffusion in Carbonated Water-Ethanol Mixtures by Molecular Dynamics Simulations.

The diffusion of carbon dioxide (CO2) and ethanol (EtOH) is a fundamental transport process behind the formation and growth of CO2 bubbles in sparkling beverages and the release of organoleptic compounds at the liquid free surface. In the present study, CO2 and EtOH diffusion coefficients are computed from molecular dynamics (MD) simulations and compared with experimental values derived from the Stokes-Einstein (SE) relation on the basis of viscometry experiments and hydrodynamic radii deduced from former nuclear magnetic resonance (NMR) measurements. These diffusion coefficients steadily increase with temperature and decrease as the concentration of ethanol rises. The agreement between theory and experiment is suitable for CO2. Theoretical EtOH diffusion coefficients tend to overestimate slightly experimental values, although the agreement can be improved by changing the hydrodynamic radius used to evaluate experimental diffusion coefficients. This apparent disagreement should not rely on limitations of the MD simulations nor on the approximations made to evaluate theoretical diffusion coefficients. Improvement of the molecular models, as well as additional NMR measurements on sparkling beverages at several temperatures and ethanol concentrations, would help solve this issue.

Involvement of capsaicin-sensitive nerves in the initiation of swallowing evoked by carbonated water in anesthetized rats.

Capsaicin powerfully evokes the swallowing reflex and is a known therapeutic agent for improving dysphagia and preventing aspiration pneumonia. However, the role of capsaicin-sensitive nerves in the initiation of swallowing evoked by various natural stimuli remains unclear. To explore this question, we blocked laryngeal capsaicin-sensitive nerves following the coapplication of QX-314 and capsaicin (QX/Cap), and investigated the effects on swallowing evoked by mechanical and chemical stimulation in anesthetized rats. Swallows were evoked by capsaicin, carbonated water (CW), distilled water (DW), and punctate mechanical stimulation using von Frey filaments applied topically to the larynx. Swallows were documented by recording electromyographic activation of the suprahyoid and thyrohyoid muscles. The initiation of swallowing by capsaicin was strongly suppressed at 5 min following QX/Cap treatment and returned in a time-dependent manner. CW-evoked swallows at 5 min following QX/Cap treatment were significantly diminished compared with before and 30 min after treatment. In contrast, DW-evoked and mechanically evoked swallows were unchanged by QX/Cap treatment. Furthermore, CW-evoked swallows were virtually abolished by transection of the superior laryngeal nerves and significantly decreased by the topical application of acid-sensing ion channel-3 (ASIC3) inhibitor APETx2, but they were not affected by the nonselective transient receptor potential channel inhibitor ruthenium red or the ASIC1 inhibitor mambalgin-1. Taken together, we speculate that capsaicin-sensitive nerves play an important role in the initiation of CW-evoked swallows.NEW & NOTEWORTHY The initiation of swallowing evoked by laryngeal capsaicin and carbonated water application was diminished by the coapplication of QX-314 and capsaicin. Carbonated water-evoked swallows were also abolished by transection of the superior laryngeal nerves and were inhibited by the acid-sensing ion channel-3 inhibitor. Capsaicin-sensitive nerves are involved in the initiation of carbonated water-evoked swallows.

An efficient utilization of chromium-containing vanadium tailings: Extraction of chromium by soda roasting-water leaching and preparation of chromium oxide.

Chromium-containing vanadium tailings (CCVT), an industrial waste, were utilized to extract chromium efficiently by soda roasting-water leaching process and for the preparation of highly pure chromium oxide. The effect of extraction of chromium under different roasting and leaching conditions were analyzed using X-ray diffraction (XRD) and scanning electron microscopy (SEM). The maximum chromium extraction rate of 91.51% was obtained when soda (Na2CO3) and CCVT were mixed in a molar ratio (n (Na2CO3)/n (Cr2O3)) of 8, roasted at 900 °C and maintained for 120 min. Then, the roasted product was leached in water at 60 °C for 60 min with a liquid-solid mass ratio (L/S) of 10. During soda roasting, the chromium-containing phase (Fe0.6Cr0.4)2O3 combines with Na2CO3 to form Na2CrO4, which was then transferred into the leaching liquid, post water leaching. The by-products such as NaFeTiO4, Na2CaSiO4, and Na0.68Fe0.68Si0.32O2 were left in the leaching residue which was called chromium tailings (CT). 87.40% chromium oxide was recovered from the unpurified leaching liquid after reduction and precipitation by adding Na2S, followed by roasting the deposit. This process not only relieved the potential threat of the industrial waste CCVT to the environment but also realized the recovery of the valuable element chromium.

Impact of radon and combinatory radon/carbon dioxide spa on pain and hypertension: Results from the explorative RAD-ON01 study.

OBJECTIVES: Therapies with low doses of radon have beneficial effects on patients suffering from chronic painful degenerative and inflammatory diseases. We already showed that this is accompanied by systemic immune modulations. We here focus on pain-reducing effects of very low doses of radon by adding carbon dioxide water and its impact on heart rate variability (HRV), blood pressure and free radicals. METHODS: 97 of 103 patients receiving radon spa (1.200 Bq/l at 34 °C or 600 Bq/l, 1 g/l CO2 at 34 °C) were monitored before and at three different time points after therapy. Individual pain perception was analyzed and the capability to process radicals. At each time point, the hypertensive patients (n = 46) were examined over 24 h for blood pressure and HRV. RESULTS: Long-term pain reduction was observed in the majority of patients. A modulation of superoxide dismutase was identified, presumably representing a priming effect for lowering radiation stress. Further, lowering of blood pressure, especially in those patients who additionally received carbon dioxide, was seen. Radon did in particular impact on HRV implying lasting relaxation effects. CONCLUSION: Radon/carbon dioxide spa efficiently reduces pain. In particular, patients simultaneously suffering from painful and cardiovascular diseases should be treated by combination of radon and CO2.

Reduction of FENO by tap water and carbonated water mouthwashes: magnitude and time course.

Fractional exhaled nitric oxide (FENO) assesses eosinophilic inflammation of the airways, but FENO values are also influenced by oral nitric oxide (NO). The aim of this pilot study was to measure FENO and compare the effect of two different mouthwashes on FENO and analyse the duration of the effect. FENO was measured in 12 randomized volunteers (healthy or asthmatic subjects) with a NIOX VERO® analyser at an expiratory flow rate of 50 mL/s. After a baseline measurement, a mouthwash was performed either with tap water or carbonated water and was measured during 20 min in 2 min intervals. The procedure was repeated with the other mouthwash. We found that both mouthwashes reduced FENO immediately at the beginning compared to the baseline (p < .001). The carbonated water mouthwash effect lasted 12 min (p ranging from <0.001 to <0.05). The tap water mouthwash reduced FENO statistically significantly only for 2 min compared with the baseline. We conclude that a single carbonated water mouthwash can significantly reduce the oropharyngeal NO contribution during a 12 min time interval.

Influence of mouthwashes on extended exhaled nitric oxide (FENO) analysis.

Fractional exhaled nitric oxide (FENO) is used to assess eosinophilic inflammation of the airways. FENO values are influenced by the expiratory flow rate and orally produced NO. We measured FENO at four different expiratory flow levels after two different mouthwashes: tap water and carbonated water. Further, we compared the alveolar NO concentration (CANO), maximum airway NO flux (J'awNO) and airway NO diffusion (DawNO) after these two mouthwashes. FENO was measured in 30 volunteers (healthy or asthmatic) with a chemiluminescence NO-analyser at flow rates of 30, 50, 100 and 300 mL/s. A mouthwash was performed before the measurement at every flow rate. The carbonated water mouthwash significantly reduced FENO compared to the tap water mouthwash at all expiratory flows: 50 mL/s (p < .001), 30 mL/s (p = .001), 100 mL/s (p < .001) and 300 mL/s (p = .004). J'awNO was also significantly reduced (p = .017), however, there were no significant differences in CANO and DawNO. In conclusion, a carbonated water mouthwash can significantly reduce oropharyngeal NO compared to a tap water mouthwash at expiratory flows of 30-300 mL/s without affecting the CANO and DawNO. Therefore, mouthwashes need to be taken into account when comparing FENO results.

More than a drink: A rare anaphylactic reaction to sparkling water.

Anaphylaxis is a potentially life threatening, type I hypersensitivity reaction which can occur within seconds to minutes after exposure to an allergen. Sulfites have been implicated in causing such reactions with symptoms ranging from mild to potentially life threatening. Here we present a patient who had an anaphylactic reaction secondary to exposure to sulfites found in sparkling water.

Bathing in carbon dioxide-enriched water alters protein expression in keratinocytes of skin tissue in rats.

Beneficial effects of balneotherapy using naturally occurring carbonated water (CO2 enriched) have been known since the Middle Ages. Although this therapy is clinically applied for peripheral artery disease and skin disorder, the underlying mechanisms are not fully elucidated.Under controlled conditions, rats were bathed in either CO2-enriched water (CO2 content 1200 mg/L) or tap water, both at 37 °C, for 10 min daily over 4 weeks. Proliferation activity was assessed by Ki67 immunohistochemistry of the epidermis of the abdomen. The capillary density was assessed by immunodetection of isolectin-positive cells. Using cryo-fixed abdominal skin epidermis, follicle cells and stroma tissue containing capillaries were separately isolated by means of laser microdissection and subjected to proteomic analysis using label-free technique. Differentially expressed proteins were validated by immunohistochemistry.Proliferation activity of keratinocytes was not significantly different in the epidermis after bathing in CO2-enriched water, and also, capillary density did not change. Proteomic analysis revealed up to 36 significantly regulated proteins in the analyzed tissue. Based on the best expression profiles, ten proteins were selected for immunohistochemical validation. Only one protein, far upstream element binding protein 2 (FUBP2), was similarly downregulated in the epidermis after bathing in CO2-enriched water with both techniques. Low FUBP2 expression was associated with low c-Myc immune-expression in keratinocytes.Long-term bathing in CO2-enriched water showed a cellular protein response of epithelial cells in the epidermis which was detectable by two different methods. However, differences in proliferation activity or capillary density were not detected in the normal skin.