WO1991011196A1

WO1991011196A1 - A method of treating a mammal with a biologically active compound

Info

Publication number: WO1991011196A1
Application number: PCT/DK1991/000021
Authority: WO
Inventors: Henrik Christensen; Søren SUSGAARD; Søren LAURBERG; Troels Torp Andreassen
Original assignee: Novo Nordisk AS
Current assignee: Novo Nordisk AS
Priority date: 1990-02-02
Filing date: 1991-01-25
Publication date: 1991-08-08
Anticipated expiration: 1992-08-02
Also published as: IE910353A1; PT96652A; AU7318691A

Abstract

A method of reducing the incidence of post-surgical problems in a mammal wherein growth hormone is administered to the individual for a period before and optionally after the operation such as an operation wherein an anastomosis is formed, a joint replacement or a skin grafting is disclosed. The administration of growth hormone increases the strength of an anastomosis and thus reduces the incidence of leakage or rupture and the mortality. The mammal to be treated is preferably a human being, and the growth hormone is preferably human growth hormone.

Description

TITLE

A Method of Treating a Mammal with a Biologically Active Com¬ pound.

The present invention relates in its broadest aspect to a method of treating a mammal with growth hormone before an operation in order to reduce the time of confinement to bed and the incidence of post-surgical problems.

The present invention relates to a method of reducing the incidence of post-surgical problems in a mammal, a method of treating a mammal with growth hormone, the use of human growth hormone for the preparation of a pharmaceutical pre¬ paration for reducing the incidence of post-surgical problems in a mammal, a pharmaceutical preparation for reducing the incidence of post-surgical problems in a mammal, and a method for the preparation thereof.

BACKGROUND OF THE INVENTION

In colorectal resections anastomotic leakage remains the most important cause of mortality (Goligher, J.C., Graham, N.G., and DeDombal, F.T.: Anastomotic Dehiscence after Anterior Re- section of Rectum and Sigmoid in Br.J.Surg. 1970; 57; 109- 18, and Debas, H.T., and Thomson, F.B.: A Critical Review of Colectomy with Anastomosis in Surg.Gynecol.Obstet. 1972; 135; 747-52). The frequency of clinical leakes varies from .1 to 30 per cent and increases the mortality rate from 7 per cent in patients with anastomoses free of leakage to 22 per cent in patients undergoing leakage (Ravo, B. : Colorectal Anastomotic Healing and intra Colonic Bypass Procedure in Ravo B. , Khubchandani IT, eds. "The surgical Clinics of North America; Philadelphia: .B. Saunders Company 1988; 1267-94 and Morgenstern, L. , Yamakawa, T. , Ben-Shoshan, M. and Lippmann, H. : Anastomotic Leakage after low Anterior Anastomosis in Am.J.Surg. 1972; 123; 104-9). Great efford has been spent trying to explain healing pro¬ cesses.

Thus, a number of factors are known to influence the healing processes during the rapid scar formation (Dunphy, J.E.: The Cut Gut in Am. .Surg. 1970; 19; 1-8, Foster, M.E. and Leaper, D.J. : Tha Alimentary Tract in Bucknall, T.E., Ellis. H. , eds. Wound Healing for Surgeons; London: Baillere Trudall 1984; 161-79 and Schrock, T.R., Deveny, C.W. and Dunphy; J.E.: Factors Contributing to Leakage of Colonic Anastomoses in Ann.Surg. 1973; 177; 513-18). The connective tissue macromolecule collagen and the balance between lysis and synthesis of collagen plays a central role in wound healing (Cronin, K. , Jackson and D.S., Dunphy, J.E.: Changing Bursting Strength and Collagen Content of The Healing Colon in Surg.Gyn.Obst. 1968; 126; 747-53, Stromberg B.V. and Klein L.R. : Collagen Formation during the Healing of Colonic Anastomoses in Dis Colon Rectum 1982; 25; 301-4, Wise, L. , McAlister, W. , Stein, T. and Schnuk, P.: Studies on the Healing of Anastomoses of Small and Large Intestine in Surg.Gynec.Obst. 1975; 141; 189-94 and Hendriks, T. , Vereecken, T.H.L.B., Hesp, W.L.E.M., Schillings, P.H.M. and deBoer, H.H.M. : Loss of Collagen from Experimental Intestinal Anastomoses: Early Events in Exper.Molec.Path 1985; 42; 411- 18) . The fibroblast is the main producer of collagen and is stimulated by growth hormone in vitro (Clemmons, D.R. and Shaw, D.S.: Purification and Biological Properties of Fibroblast Somatomedin in J.Biol.Chem. 1986; 261; 10293-98) .

In spite of all these investigations there has until now been no disclosure or indication of how to reduce the incidence of leakage or rupture of anastomoses.

It is an object of the present invention to provide a means to reduce the incidence of anastomotic leakage in order to reduce the risk in connection with anastomoses. When inserting prostheses having high loading, especially hip joints, knee joints, or ankle joints is is crucial to have a very quick inset of healing and a rapid development of strength as it is essential for the patients to avoid confinement to bed as early as possible in order to avoid thrombosis and to ensure a proper healing in order to avoid loosening of such a joint. The same applies when repositioning complicated fractures of a bone, e.g. after stretching, e.g. if a first reposition has failed. Such fractures will heal more quickly after administration of hGH which is of special importance for elderly people.

In severe burns it is often necessary to carry out a skin grafting to ensure a healing. In such cases a quick onset of healing and proper healing are crucial for the succes.

In accordance with the present invention it has now surpri¬ singly been found that the administration of human growth hormone to a patient who is to have an anastomosis reduces the rise of leakage or rupture, and thus the mortality, to a very high degree. Furthermore, such administration of growth hormone ensures a very quick inset of healing and a rapid development of strength when inserting prostheses such as hip joints, knee joints, or ankle joints and a quick onset and proper healing of burns after skin grafting.

BRIEF DESCRIPTION OF THE INVENTION

The present invention relates, according to a first aspect thereof, to a method of reducing the incidence of post- surgical problems in a mammal wherein growth hormone is ad¬ ministered to the individual for a period before and optionally after the operation.

According to the invention,growth hormone may be administered to a mammal for a period before and optionally after an operation wherein an anastomosis is formed, a joint replacement or a skin grafting.

In accordance with a preferred aspect of the invention growth hormone is administered to an individual for a period before and optionally after an operation wherein the anastomosis is formed. Such administration may be before and optionally after an operation of colon or rectum, preferably an operation of a polypus or cancer.

According to a preferred aspect of the invention, the mammal to be treated is a human being, and wherein the growth hormone is human growth hormone.

According to a third aspect, the invention provides the use of a growth hormone for the preparation of a pharmaceutical preparation for treating a mammal for a period before and optionally after an operation wherein an anastomosis is for¬ med.

In accordance with a preferred aspect of the invention, human growth hormone is used for the preparation of a pharma¬ ceutical preparation for treating a human being for a period before and after an operation. Such operation may be an operation wherein an anastomosis is formed, a joint replacement, a reposition of a fractured bone or a skin grafting.

According to a more preferred aspect of the invention, the preparation may be used when carrying out an operation of colon or rectum, preferably an operation of a polypus or cancer.

According to a further aspect, the invention provides a phar¬ maceutical composition for reducing the incidence of post- surgical problems in a mammal comprising growth hormone op¬ tionally in combination with a pharmaceutical acceptable carrier and/or excipients for lyophilizing. Post-surgical problems which may be treated using the preparation of the invention are e.g. leakage or rupture of an anastomosis after an operation wherein an anastomosis is formed or loosening of a replaced joint or non-healing after a skin grafting operation.

According to a still further aspect, the invention provides a method for preparing a pharmaceutical composition for redu¬ cing the incidence of post-surgical problems in a mammal comprising mixing human growth hormone, optionally in combination with a pharmaceutical acceptable carrier, with excipients for lyophilizing and lyophilizing the resulting mixture. The human growth hormone is preferably pure so as to yield a single major band on a polyacryl amide gel.

The invention may be worked using any growth hormone such as human growth hormone or an animal growth hormone such as bovine, porcine, equine, canine, avian or an ovine growth hormone. However, the growth hormone used is preferably the one corresponding to the species of the individual to be treated, e.g. human growth hormone for treating human beings etc.

The administration of growth hormone according to the inven¬ tion may be carried out by subcutaneous, intravenous or intramuscular injection or by intravenous infusion of a solution comprising the growth hormone.

The dosis may vary from 0.1 to 4 mg GH/kg bw per day pre¬ ferably from 1 to 3 mg GH/kg bw per day, more preferred about 2 mg GH/kg bw per day which is preferably given in divided doses two times a day

The administration of growth hormone according to the inven- tion may be carried out from 10 days prior to the operation to 10 days after the operation, preferably for a period form

7 days prior to the operation and 7 days after. In some cases the period may be from 4 days prior to 4 days^" after the operation.

The total dosis given to a human being for such a period of 14 days may vary from 50 to 150 IU growth hormone.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention is explained more in detail with reference to the drawings in which

Fig.l shows a graphical representation of the changes of the initial body weight in rats undergoing a left colonic resection,

fig.2 shows a graphical representation of the bursting pressure in vivo of left colonic anastomoses, and

Fig.3 shows a graphical representation of the bursting wall tension in vivo of left colonic anastomoses.

DETAILED DESCRIPTION OF THE INVENTION

The invention is explained more in detail in the below Exam¬ ple illustrating the invention. The examples presents embodi¬ ments of the invention but are, however, not to be considered as limiting the scope of the invention being defined by the appending claims.

EXAMPLE 1

Seventy-two 3 months old female Wistar rats (Møllegaard, Denmark) weighing 227 ± 1 g (mean ±SEM) were randomized into 2 groups for treatment with saline (controls) or 2.0 mg biosynthetic human growth hormone (b-hGH) per body weight per day (specific activity: 1 mg = 3 I.U., Novo-Nordisk A/S, Gentofte, Denmark) . All injections were started 7 days before operation and continued until sacrifice. The isotonic growth hormone solution was injected subcutaneously in the nape of the neck in two daily doses and the control animals were injected with a corresponding dose of saline. The animals were weighed at the start of experiment, on the day of operation and on the day of the testing procedure. They had free access to normal pellet food (Altromin diet 1324, CHr. Pedersen Ltd., Ringsted, Denmark) and tap water. A survey on the experiments appear from the below Table I.

Table I. Number of rats included in the study. The animals received daily injections of saline (controls) or 2.0 mg biosynthetic human growth hormone (b-hGH) starting 7 days before a colonic anastomosis was performed and continued until the day of testing. Excluded animals in parentheses.

Injections Saline b-hGH

Day after surgery

2 12 11 (l)

4 10 (2) 10 (2) 6 10 (2) 12

Total 34 (4) 33 (3)

Operation: Under general anaesthesia (intraperitoneal injection of Pentobarbital 50 mg/ml; 0.1 ml/100 mg body weight) an abdominal midline incision was made. By a clean procedure a 1.0 cm segment of the left colon was resected 3 cm proximal to the rectal -peritoneal reflection, preserving marginal vessels. Faecal content was milked out around the anastomotic site. A standardized end-to-end anastomosis was made with a single layer of eight interrupted inverting full- thickness 6-0 polypropylene sutures (Prolene R, Ethicon Ltd.) .

Testing procedure: The anastomoses were tested in vivo and in sity on the 2nd, 4th, or 6th postoperative day. A re- laparotomy was performed under general anaesthesia as described previously. Adhesions were evaluated according to an arbitrary grading scale (Uden, P., Blomquist, P., Jiborn, H., Zederfeldt, B. , in Acta Chir Scand 1988: 154: 127-32). The results appear from the below Table II.

Table II. Adhesions around left colonic anastomoses in rats injected with 2.0 mg biosynthetic human growth hormone (b- hGH) or saline (controls) for 7 days preoperatively and until sacrifice on the 2nd, 4th, and 6th postoperative day.

Adhesion grade 1 2 3 4

Number of animals saline 0 2nd day b-hGH

saline 4th day b-hGH

saline 6th day b-hGH

The amount of perianastomotic adhesions are graded by the arbitrade scale as 1: no adhesions; 2: minor adhesions, 3: more than half the anastomosis area covered with adhesions; 4: anastomosis completely covered with adhesions.

The perianastomotic adhesions were removed carefully. The colon was opened 3 cm proximal to the anastomosis and the faecal content was gentely milked out. Through the colostomia a double-barrelled pressure catheter was placed at the anastomosis site. A suture was tightened around the colon and the catheter 2.0 cam proximal to, and around the colon 2.0 cm distal to the anastomosis. Isotonic saline (21°C) was insufflated through the catheter at a constant rate of 2.5 ml/min by means of a perfusor (Holm & Halby until rupture. simultaneously, the intraluminal pressure of the isolated colonic segment was recorded via a pressure-transducer and a differential-amplifier on an X-Y recorder. The bursting pressure (BP) was the maximum pressure. The site of leakage was noted.

The tested segment kept its cylindrical form during inflation except for a small restriction at the anastomosis. By regarding the intestinal segment as a cylinder, the radius of the anastomosis at rupture could be calculated from the amount of insufflated saline.

The bursting wall tension (BWT) could then be calculated according to La Place's law (BWT = BP x radius). All anastomoses were tested with sutures in situ to imitate the clinical situation and in order not to damage the anastomosis by removal of sutures.

The results appear from the below Table III .

Table III. The bursting pressure (BP) and the bursting wall tension (BWT) and the bursting radius (r) of left colonic anastomoses on the 2nd, 4th, and 6th postoperative day in rats injected with saline (controls) or 2.0 mg biosynthetic human growth hormone (b-hGH) per kg per day.

6th day b-hGH 216 ± 10.2 127.3 ± 7.2 5.8 ± 0.2 Mean values ±SEM. *: 2p < 0.05; ***: 2p < 0.005, compared to saline controls.

Exclusion of animals: Three animals with an anastomotic abscess (1 control, 2 b-hGH animals) were excluded. One control animal was excluded due to a lesion of the anastomosis at the relaparotomy. Three animals (2 controls, 1 b-hGH) died from anaesthetic complications.

Bursting strength of normal colon: nine rats of the same strain, age, sex and weight (214 ± 5 g; mean ±SEM) were tested by the same procedure to determine the bursting pressure and bursting wall tension of normal left colon. After testing the animals were sacrificed with an intracardial overdose of barbiturate.

Statistical methods: Comparison of breaking sites between the two groups was analyzed by the Fischer's test. Analysis of the other data was performed by the non-parametric Mann- Whitney two-sample test. 2P values < 0.05 were considered significant. RESULTS

The figures illustrate the results. Fig.l shows the changes of body weight in per cent of the initial body weight (g) in female rats undergoing a left colonic resection. "." 5 represents animals injected with saline (controls) , and "0" represents animals injected with 2.0 mg biosynthetic human growth hormone (b-hGH)/kg body weight/day.

S: Start of injections; Op: Operation; Bars represent 1 SEM; ***: 2p < 0.005, as compared with controls.

10 Fig. 2 shows the bursting pressure (BP) in vivo of left colonic anastomoses on the 2nd, 4th, and 6th postoperative day in rats injected with 2.0 mg biosynthetic human growth hormone (b-hGH)/kg body weight/day or saline (controls) . Op: Operation. Bars represent 1 SEM; *: 2p < 0.05; ***: 2p <

150.005, as compared with controls.

Fig.3 shows the bursting wall tension (BWT) in vivo of left colonic anastomoses on the 2nd, 4th, and 6th postoperative day in rats injected with 2.0 mg biosynthetic human growth hormone (b-hGH)/kg body weight/day or saline (controls) . n:

20 number of rats; Op: Operation; Bars represent 1 SEM; *: 2p < 0.05; ***: 2p < 0.005, as compared with controls.

Body weights (cf Figure 1)."

After 7 days preoperative injections the body weights of the b-hGH injected animals were increased by 14% when compared to

25 the control group. In both groups the body weights were decreased by 3% on the 2nd postoperative day. On the 4th and 6th postoperative day the body weights of the controls were 97% of the weight at the time of operation, whereas the b- hGH injected animals regained their weight at the time of

30 operation on the 4. postoperative day.

Bursting strength of the colonic anastomoses

The results for colonic anastonotic strength in rats are shown in detail in Table 2 and Figures 2 and 3. The bursting pressure was significantly increased by 104% in the b-hGH

35 injected group on the 2nd postoperative day (2p < 0.005) and by 232% on the 4th postoperative day (2p < 0.05) when compared to the saline injected control group. The bursting wall tension was significantly increased by 133% in the b-hGH injected group on the 2nd postoperative day (2p < 0.005) and by 278% on the 4th postoperative day when compared to the saline injected control group. There was no difference between the two groups on the 6th postoperative day. Bursting strength of normal left colon. The bursting pressure of intact left colon segments in the 9 nonoperated animals was 255 ± 1 Pa-10² (mean ±SEM). The colonic anastomotic bursting pressure on the 6th postoperative day was 84% for the saline treated group and 85% for the b-hGH treated group, compared to the bursting pressure of intact colon. The bursting wall tension of intact colon was 121 ± 14 N/m (mean ±SEM) . The anastomotic bursting wall tension on the 6th postoperative day for the saline treated group was 99% and for the b-hGH treated group was 105%, compared to the bursting wall tension of intact colon. Breaking pattern

On the 2nd and 4th postoperative day all anastomoses in both groups broke at the anastomotic line. On the 6th postoperative day a difference of breaking pattern was found between the groups. In the control group 5 anastomoses broke at the incisional line and 5 broke in a zone adjacent to the incisional line distally to the anastomoses. In the b-hGH injected group 11 out of 12 anastomoses (92%) broke distally to the incisional line. Adhesions No difference was found in the amount of perianastomotic adhesions between the control group and the b-hGH injected group (cf Table 3) .

Bursting radius of the colonic anastomoses. The radii of the anastomoses at the time of rupture was significantly larger in the b-hGH injected animals on the 2nd and 4th postoperative day (2p < 0.05) when compared to the controls (cf Table 2) . There was no difference on the 6th postoperative day. Bursting radius of intact colon.

The radius of intact colon at breakage was 5-3 ± 0.1 mm (mean ±SEM) .

DISCUSSION

The experiments show that growth hormone, given as biosynthetic human growth hormone, increases the bursting pressure and the bursting wall tension of left colon anastomoses in rats which have received injections of growth hormone for 7 days before colonic resection. The findings of a two-fold increase of the bursting pressure and the bursting wall tension on the 2nd postoperative day and a three-fold increase on the 4th postoperative day indicates that growth hormone can substantially influence the early phases of the healing process of the colon. The bursting strength of the anastomoses at the 6th postoperative day in both groups were at the same level as the intact unoperated colonic strength and this may explain why no difference was found on this day. However, it is to be noted that on the 6th postoperative day more than 92% in the b-hGH group disrupted outside the colonic anastomosis.

Since nearly all colonic segments disrupted outside the anastomotic line in the b-hGH injected animals on the 6th postoperative day it is believed that the stimulating effect of growth hormone is more related to healing of the anastomosis than to a generalized increased strength of the colonic intestine wall.

Bursting strength, expressed as bursting pressure or bursting wall tension, is a measure of the wall resistance to increases in the intraluminal pressure and has been widely used, because it is considered to reflect the physiologic strain in the intestine more accurately than breaking strength.

The present data suggest that treatment with biosynthetic human growth hormone might reduce the risk of leak from colonic anastomoses. The leak rate in a clinical context is a failure of healing with disruption of a part of the suture line, whereas the colonic bursting strength is an objective measurement of the healing anastomosis. Nevertheless, it is generally suggested that factors improving the anastomotic strength also would be expected to reduce the risk of a clinical leak.

EXAMPLE 2

THE INFLUENCE OF THE DOSIS OF GROWTH HORMONE ON THE HEALING OF COLONIC ANASTOMOSES

The aim of the study was to investigate the effect of different doses of biosynthetic human growth hormone (b-hGH) on the anastomotic strength in healing experimental anastomoses. The experiments were carried out as explained in Example 1 using 12 rats in each group. The bursting pressure was determined as described in Example 1. The collagen contents (the hydroxyproline contents HPC) was determined as follows: 6 mm samples collected from a 6 cm segment comprising the anastomosis were freeze-dried. The samples were weighed (Dry weight (DW) ) and defatted by acetone for 72 hours with one change of acetone after 48 hours, freeze dried and weighed again, (defatted dry weight (DDW) ) . The hydroxyproline contents was determined according to Woessner (in Hall DA, ed. The Methodology of Connective Tissue Research. Oxford: Joynson-Bruwers, 1976, 227-33) .

Growth hormone was given in four different doses, viz. .125, .5, 2.0, and 8.0 g/kg body weight/day. The treatment was initiated 4 days before operation and continued until 4th post-operative day (test) . Saline injected animals were used as controls. The results appear from the below Table IV. TABLE IV.

The bursting pressure (BP) and collagen contents (HPC) and the defatted dry weight (DDW) .

X ± SEM BP HPC DDW

5 (Pa-10²) (mg/sp) (mg)

Saline 20.09 ± 0.28 0.277 ± 0.016 29.03 ± 1.48 (controls)

0,125 mg 31.23 ± 0.67 0.306 ± 0.023 30.55 ± 2.17 10 bhGH/kg/day

0,5 mg 44.5 ± 1.12 0.332 ± 0.026 35.40 ± 2.41 bhGH/kg/day *

2,0 mg 56.12 ± 0.84 0.444 ± 0.016 48.66 ± 2.00 bhGH/kg/day ** ** **

158.0 mg 87.01 ± 1.11 0.448 ± 0.020 54.33 ± 3.66 bhGH/kg/day ** ** **

**: P < 0.005 *: P < 0.05

The results show that response depends on the dosis

20 EXAMPLE 3

INCREASE OF THE STRENGTH OF INTACT BONES AND HEALING TIBIAL FRACTURES IN RAT (DUE TO ADMINISTRATION OF GROWTH HORMONE) .

The aim of the study was to investigate the influence of exo¬ genous growth hormone administration on the biomechanical

25 properties of healing diaphyseal fractures in the rat. In 90- day-old female Wistar rats a standardized tibial fracture was produced in the right tibia and non-rigid fixation was esta¬ blished using a K-wire as intramedullary nail. Two mg of bio¬ synthetic human growth hormone (b-hGH) per kg body weight per

30 day was given in 2 daily injections starting one week before fracture and continuing until testing. The control groups were injected with saline. After 40 days of fracture healing, the rats were killed and the fractured and the corresponding non-fractured bones were tested in a materials testing machi¬ ne using a destructive three-point bending procedure. The results appear from the below Table V.

Table V

Differences noted between growth hormone injected and con¬ trols after 40 days treating:

In the growth hormone injected animals, maximun load and stiffness of the intact bones had increased 40 days post fracture compared to the saline injected controls, but there was no difference in terms of stress values, modulus of ela¬ sticity or normalized energy absorption.

In conclusion, growth hormone stimulates the mechanical strength development in healing diaphyseal fractures in the rat, when a total dose of 2 mg/kg BW/day is given in 2 daily injections. An increased strength observed in the non-frac¬ tured bones seems to be a quantitative phenomenon. EXAMPLE 4

THE INFLUENCE OF THE DOSIS OF GROWTH HORMONE ON FRACTURE HEALING IN THE RAT

Growth hormone stimulates the proliferation of chondrocytes in vivo and in vitro. Growth hormone also stimulates weight gain and longitudinal bone growth depending on the dose and freqency of administration. Experimental fractures of long bones heal through a stage of cartilaginous callus if the mechanical conditions are not absolutely stable. The aim of the study was to investigate the effect of different doses of biosynthetic human growth hormone (b-hGH) on the mechanical strength development in healing experimental fractures. In 90-day-old female Wistar rats, a closed fracture was pro¬ duced by three-point bending 2 mm above the tibio-fibular junction in the right tibia. Closed medullary nailing was performed, and the bones were left to heal for 40 days. The rats were randomized into 6 groups: no injections, 0.9% NaCl (volume corresponding to b-hGH-treated groups), 0.08, 0.4, 2.0 and 10 mg b-hGH/kg/day given in 2 daily doses, starting one week prior to fracture and continuing until testing. Bio- mechanical testing was carried out in a materials testing machine by a three-point bending procedure. Load and deforma¬ tion was recorded continously, and maximum load, stiffness and energy absorption were calculated. The results appear from the below Table VI.

Table VI

Increased maximum load and stiffness of fractures treated with b-hGH in doses of 2 mg/kg/day and 10 mg/kg/day after 40 days healing as compared with controls: Treatment n Maximun load Stiffness (N) (N/mm)

51) no injections 13 37.2 ± 6.5 171. 0 ± 31. 0

2) 0.9%NaCl 14 30.2 ± 4 .9 138. 0 ± 24 .2

3) b-hGH 0.08 mg/kg/day 10 35. 6 ± 8 .2 166. 0 ± 39 . 3

4) b-hGH 0.4 mg/kg/day 15 34 .9 ± 5.8 179.7 ± 30.5

5) b-hGH 2.0 mg/kg/day 10 55.3 ± 10.2 219.9 ± 31.8 106) b-hGH 10 mg/kg/day 13 69.3 ± 8.4^** 323.0 ± 30.0^*'

Mean values ± SEM; 2p < 0.05; 2p < 0.01

Conclusion: Biosynthetic human growth hormone administered twice a day accelerates the mechanical strength development 15 in healing rat tibial fractures when given subcutaneously in doses of 2 mg/kg/day and 10 mg/kg/day.

Claims

1. A method of reducing the incidence of post-surgical problems in a mammal wherein growth hormone is administered to the individual for a period before and optionally after

5 the operation.

2. A method as claimed in claim 1 wherein the growth hormone is administered to a mammal for a period before and optionally after an operation wherein an anastomosis is formed, a joint replacement or a skin grafting.

103. A method as claimed in claim 2 wherein growth hormone is administered to an individual for a period before and after an operation wherein an anastomosis is formed.

4. A method as claimed in claim 3, wherein the growth hor¬ mone is administered for a period before and after an

15 operation of colon or rectum.

5. A method as claimed in claim 4, wherein the operation concerns a polypus or cancer.

6. A method as claimed in any of claims 1 to 5 wherein the mammal to be treated is a human being, and wherein the growth

20 hormone is human growth hormone.

7. Use of a growth hormone for the preparation of a pharma¬ ceutical preparation for treating a mammal for a period before and optionally after an operation.

8. Use of a growth hormone as claimed in claim 7, wherein 25 human growth hormone is used for the preparation of a pharma¬ ceutical preparation for treating a human being for a period before and optionally after an operation wherein an anastomosis is formed, a joint replacement or a skin grafting. 9. A pharmaceutical composition for reducing the incidence of post-surgical problems in a mammal comprising growth hormone optionally in combination with a pharmaceutical acceptable carrier and/or excipients for lyophilizing.

510. A method for preparing a pharmaceutical composition for reducing the incidence of post-surgical problems in a mammal comprising mixing growth hormone, optionally in combination with a pharmaceutical acceptable carrier, with excipients for lyophilizing and lyophilizing the resulting mixture.