NON-SHIVERING CRYOTHERMOGENESIS METHOD FOR REDUCING ADIPOSE TISSUES

Abstract

The present disclosure refers to a non-shivering cryothermogenesis method for locally reducing adipose tissue of a subject's body, including: (a) applying at least one applicator to the subject's skin; (b) applying at least one additional applicator to a local region wherein adipose tissue reduction is desired; (c) the applicator maintaining skin superficial temperature between 4 C. and 12 C.; and (d) applying the applicators for a total period of time between 20 and 60 min.

Claims

1. A non-shivering cryothermogenesis method for locally reducing adipose tissue of a subject's body, comprising: (a) applying at least one cooling applicator to the subject's skin to region wherein subcutaneous adipocytes tissue is rich in brown adipocytes and/or in white adipose tissue (WAT) containing brown adipocytes; (b) applying at least one additional cooling applicator to a local region wherein adipose tissue reduction is desired, if said local region differs from step (a); (c) said cooling applicator maintaining skin superficial temperature between 4 C. and 12 C.; and (d) applying said cooling applicators for a total period of time comprised between 20 and 60 min.

2. The method of claim 1, wherein said cooling applicator applies a temperature comprised between 15 C. to 5 C.

3. The method of claim 1, wherein said time period of step d) is divided in two or three subperiods, each of said subperiods being between 10 and 20 minutes.

4. The method of claim 1, wherein said three subperiods consist in: i. applying a temperature of 10 C. during 10 minutes; and ii. applying a temperature of 8 C. during 10 minutes; and iii. applying a temperature of 7 C. during 20 minutes.

5. The method of claim 1, wherein said local region is selected from the stomach, love handles, inner thighs, inner knees, butt, inner arms, chin, breast area, back and neck.

6. The method of claim 1, wherein 5 to 14 of said cooling applicators are applied to said subject's skin.

7. The method of claim 1, wherein said cooling applicators are applied to the subject's skin with a pressure comprised between 50 kPa and 300 kPa.

8. The method of claim 7, wherein said pressure is comprised between 50 and 150 kPa when the thickness of the skin of a subject is inferior or equal to 3 cm, and is comprised between 151 and 300 kPa when the thickness of the skin of a subject is superior to 3 cm.

9. A cryothermogenesis method for reducing adipose tissue of a subject's body, comprising the step of: (a) firstly applying a non-shivering cryothermogenesis method to a back of the subject; and then (b) secondly applying the non-shivering cryothermogenesis method to a front of the subject; the non-shivering cryothermogenesis method comprising: (i) applying at least one cooling applicator to the subject's skin to region wherein subcutaneous adipocytes tissue is rich in brown adipocytes and/or in white adipose tissue (WAT) containing brown adipocytes; (ii) applying at least one additional cooling applicator to a local region wherein adipose tissue reduction is desired, if said local region differs from step (i); (iii) said cooling applicator maintaining skin superficial temperature between 4 C. and 12 C.; and (iv) applying said cooling applicators for a total period of time comprised between 20 and 60 min.

10. A non-shivering cryothermogenesis method for locally reducing adipose tissue of a subject's body, comprising: (a) applying at least one cooling applicator to the subject's skin to region wherein subcutaneous adipocytes tissue is rich in brown adipocytes and/or in white adipose tissue (WAT) containing brown adipocytes; (b) applying at least one additional cooling applicator to a local region wherein adipose tissue reduction is desired, if said local region differs from step (a); (c) said cooling applicator maintaining skin superficial temperature between 4 C. and 12 C.; (d) applying said cooling applicators for a total period of time comprised between 20 and 60 min; and (e) repeating the method of steps (a)-(d) to the subject's skin a plurality of times until a desired reduction in the adipose tissue at said local region has been achieved, wherein said application can be repeated as soon as said adipose tissue recover their temperature of prior said step (a) or (b).

11. The method of claim 9, wherein said plurality of times are spaced of 1 hour time intervals, 12 hour time intervals or 1 day time interval.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0041] FIG. 1. Percentage change in (A) waist circumference, and (B) thigh circumference, following single abdominal applications of tissue cryotherapy (n=17; treatment duration ca 1.33 h; time following treatment, 1 h). Error bars indicate SD.

[0042] FIG. 2. Individual responses to tissue cryotherapy as a function of age in subjects receiving single applications of tissue cryotherapy (trendline added, P=0.2).

[0043] FIG. 3. Decrement in (A) waist circumference, (B) weight, and (C) BMI following three sequential daily tissue cryotherapy treatments (n=12). Error bars indicate SD.

[0044] FIG. 4. Progressive decline in waist circumference (expressed as %) in five subjects undergoing repeat tissue cryotherapy treatments over four (subject 1, M, 59 years of age), five (subjects 2, 3, 4; F, 47 years; F, 71 years; F, 52 years), or seven (subject 5, M, 51 years) days. The mean decline after 5 treatments was 6.1%.

DETAILED DESCRIPTION AND EXAMPLES

Example 1

Effect of the Cryothermogenesis Procedure on Waist Circumference

[0045] Methods

[0046] 1. Subjects and Ethical Permissions

In this retrospective study data for 28 participants (9 M/19 F), mean age 51.5 (range 33-71 years) were analyzed. All had a BMI consistent with overweight. Thermogenesis method, a non-invasive procedure, does not require formal institutional ethical approval.

[0047] 2. Tissue Cryotherapy Procedure

Treatment employed a commercial tissue cryotherapy device (FG6601-006, ADSS, Republic of China) equipped with multiple cooling probes (cooling area 208 cm). Weight, height, and waist and thigh (left) circumferences were recorded; BMI was calculated automatically using a commercial apparatus (model SC240MA, Tanita, Japan). A layer of wetted paper (glycerol-based wetting membrane, ETG-111-200, Freezefats, Republic of China) was applied to the lower back of the reclining subject, followed by symmetrical application of four to six pairwise probes accompanied by gentle suction to improve contact. Cooling temperature was set to 10 C., declining to 7 C. over 20 minutes; application duration was 40 minutes. Subjects then lay supine and the treatment was repeated on the lower abdomen, again for 40 minutes. Oral temperature, a proxy for core temperature, was recorded before and after the procedure. Local skin surface temperatures were recorded immediately after removal of the cooling probe and measurement showed that it was maintained around +8 C. and did not fall below +4 C. Weight, waist and thigh circumference, body weight, and BMI were recorded again after the procedure. 12 subjects volunteered for triple daily applications. Five participants (3 F and 2 M; aged 47, 51, 71; and 51, 57 years, respectively) volunteered for more extended daily applications, and the procedure was repeated on a daily basis for a total of four, five, or seven applications, with systematic recording as before.

[0048] 3. Statistics

Statistical analyses employed normalized means and SD. Results were evaluated using the Student t test; statistical significance was set at P0.05.

[0049] 4. Local Cooling Produces Decrements in Waist Circumference

Waist circumference was recorded before and after single applications of local abdominal tissue cooling (cryothermogenesis method) in a series of 17 subjects. As shown in FIG. 1A, there was a mean reduction of 3.0% of waist circumference following the procedure. The decline was of high statistical significance (P=<0.0001). By contrast, there was no change in thigh circumference (FIG. 1B). Core body temperature (oral) rose from mean 35.8 to mean 36.1 (+0.3 C.), but the change was not significant. Changes in weight and BMI with single applications fell short of statistical significance (not presented; see below).
As shown in FIG. 2, there was a trend towards a reduction in efficacy as a function of age. However, the trend was not of statistical significance (P=0.2): some older individuals displayed substantial reductions in waist circumference and some young subjects displayed low responses. There were no significant gender differences.

[0050] 5. Multiple Applications Produce Loss of Waist Circumference

Twelve subjects undertook subsequent treatments on days 2 and 3 under the same conditions and for the same duration as on day 1. As shown in FIG. 3, in addition to a significant decline in waist circumference (P=0.0068), highly significant reductions in both body weight (P=<0.001) and BMI (P=0.013) were observed. The extent of mean reduction in waist circumference, 4.2%, exceeded the mean reduction obtained by single applications (3.0%, P=NS versus single application).
To address a possible delay-/time-dependent effect, nine subjects receiving a single treatment underwent further measurements 5 to 10 days later but without further tissue cryotherapy. Given the small number of subjects, the extent of the reduction in waist circumference (3.9%, SD=3.09) did not achieve statistical significance versus the reduction observed immediately following single treatment (3.0%, SD=2.71), although there was trend towards an increase.
Five subjects (3 F, 2 M) volunteered for repeat daily treatments over periods of 4, 5, and 7 days. As shown in FIG. 3, repeat treatment was accompanied, in all five subjects, by a progressive loss of waist circumference (mean loss of 6.1% immediately following five treatments), to be compared against a 3% mean loss immediately following a single treatment. The difference in waist loss following four to seven treatments was significantly greater than from single treatments (P=<0.01).

[0051] 6. Biomedical Parameters

Local skin temperatures immediately beneath the cooling probe were variable within the region, but did not fall below +6 C. There was some local reddening of the skin consistent with increased local blood circulation. No superficial skin damage of any type was observed. Comprehensive blood analysis was carried out on four subjects before and 3 days after treatment. Profiles before and after treatment were unremarkable. Importantly, except as noted, there was no indication of inflammation, macrophage activation, or abnormal lipid profiles. Therefore the cryothermogenesis method is not associated with systemic inflammation or changes in blood lipid profiles. In addition, no participant reported any adverse events.

[0052] 7. Conclusion

Single applications of local abdominal tissue cooling (cryothermogenesis method) were found to produce a significant tissue reduction. All subjects responded to single treatments by a reduction in waist circumference (mean 3%) that was statistically significant (P=<0.0001). There was no evident M/F gender difference in response. There was an apparent trend towards to reduced efficacy with age; however, the trend was not statistically significant, and participants among the oldest in the group displayed responses to single treatments.

[0053] Results show further progressive loss of AT with multiple applications. A larger reduction in circumference was observed following three treatments (4.2%), and this was accompanied by decrements in both body weight (0.75%) and BMI (0.8%) that were statistically significant. The reduction in BMI indicates selective loss of AT volume. The reduction in waist circumference after three treatments (0.4%) was consistent with the decline in total body weight (0.75%).

[0054] In five subjects receiving extended rounds of treatment (four to seven), there was evidence for progressive decrements in waist circumference (6% after five treatments), arguing that multiple treatments at daily intervals produce cumulative benefits. No consistent changes in biochemical parameters or adverse events were reported for the participants in this study. These results confirm that local cooling of abdominal tissue produces selective loss of AT volume.

[0055] In contrast to earlier reports that there is no evident change in body fat immediately after treatment, the results showed significant beneficial effects within 1 h of treatment, a time-course too rapid to be explained by cell death and clearance. In addition, multiple daily treatments produced progressive losses of waist circumference, bodyweight, and BMI which is incompatible with the cell disruption. Furthermore, biochemical analysis revealed no evidence of markers of inflammation or macrophage activation in treated subjects, showing adipocyte cell volume reduction via thermogenesis, with conservation of cellular integrity. In support, complete maintenance of core body temperature (+0.3 C.) was recorded despite intense local cooling, attesting that thermogenesis is taking place. Results therefore point to energy expenditure via cold-induced fat metabolism and thermogenesis, with consequent reduction in adipocyte cell volume and AT mass.

[0056] Regarding the safety of the method according to the invention, no consistent changes in biochemical parameters were observed following treatment, including markers of inflammation. No adverse effects were reported by the participants.

Example 2

Inflammatory Biomarkers Measurements

[0057] Method

[0058] 1. Biomedical Parameters

Comprehensive blood analysis was carried out on subjects before and 3 days after the daily treatment applied daily during 5 to 10 days. All profiles before and after treatment (4 days) were unremarkable. Importantly, there was no indication of inflammation or macrophage activation.

[0059] Results

TABLE-US-00001 TABLE 1 Measure of Neopterine before and after cryothermogenesis treatment Neopterine (nmol/l) Subject 1 before 3.3 after 8 Subject 2 before 2.9 after 5.2 Subject 3 before 9 after 6.5 Subject 4 before 4 after 6.6 Subject 5 before 8.1 after 7.4

TABLE-US-00002 TABLE 2 Measure of the sedimentation rate (first hour and second hour) and C-reactive protein (CRP) before and after cryothermogenesis treatment Sedimentation rate CRP Subject a before 7 and 19 <5 after 6 and 13 <1 Subject b before 4 and 9 3 after 9 and 26 5 Subject c before 3 and 6 <5 after 3 and 6 <5 Subject d before 6 and 14 <5 after 6 and 15 2 Subject e before 2 and 6 5 after 3 and 5 <1

[0060] Conclusions

Results of table 1 and 2 showed that cryothermogenesis method is not associated with systemic inflammation as neopterine (normal value: <25 nmol/I, the sedimentation rates (normal values: first hour 1 to 20 and second hour 4 to 40) and CRP (normal value: <6 mg/L), which are well known by the skilled person, are in the range of their respective normal values. In addition, no participant reported any adverse events.

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