Collagen peptides and Joint & Bone Health Rousselot studies and literature

Improvement by nature

Joint & Bone Health

As the population ages, the drawbacks of aging are becoming more are among the leading causes of pain and disability.

Aging of the worlds population sees growth in bone and joint health problems.
apparent. Osteoarthritis and osteopenia, two of the major health concerns,

It is now well known that collagen peptides can help to maintain bone and joint health to prevent osteopenia and osteoarthritis.

According to the World Health Organization (WHO), osteopenia is used to describe individuals who have a low bone mass and some increased risk of fracture. Their bone mass is not

The main goal for screening and treating for osteopenia is to maintain bone health and prevent fractures. It has been demonstrated that an early diagnosis and treatment of osteopenia reduces fractures rates and improves life quality (3). In a population-based random study (616 women

Bones
Bones, which makes up the skeleton, are composed of cells embedded in hard intercellular material (the matrix) made of mineralized substances and collagen bers. Like all body tissues, bones are in a continuous state of ux. The mechanical integrity of the skeleton and the maintenance of appropriate mineral levels depend on a dynamic process called bone remodeling or bone turnover. Skeletal mass increases progressively during growth. The age at which bone loss starts is uncertain but, it is generally believed to be during the thirties in both sexes. Approximately 15% of bone in healthy adults is replaced by bone turnover each year. Irregular rates of bone resorption and formation leading to more bone loss than formation are a hallmark to osteopenia.

so low that they are deemed to have osteoporosis. Bone mineral T-score measure +1,0 0,0 -1,0 -2,0 -2,5 -3,0 -4,0 Normal bone density Low bone mass Presence of osteoporosis

aged 60 - 94 years followed for a median 5.6 years) conducted by Pasco and published in 2006 in Osteoporosis journal (13), it has been determinated that 73.1% of fractures occurred in women without osteoporosis (56.5% in women with osteopenia and 16.6% in women with normal Bone Mineral Density).

The bone mass is measured on a point scale, called a T score. The range dening osteopenia or low bone mass is quite large, ranging from -1 to -2.5 T-scores (that is, bone density levels that are 1 to 2.5 standard deviations below the average for young adults who have achieved normal peak bone mass), which translates into bone mass that is 1030 percent below this average level. 2500 1990 Hip fractures (103) 2000 2025 1500 2050 1000 500 0 Men Women North America Men Women Europe Men Women Russian Federation

Because life expectancy is increasing around the world, the number of elderly individuals is rising in every region. According to WHO, the number of individuals aged 65 years or over will increase to an estimated 1555 million by the year 2050. The inuence of such changes on the number and regional distribution of hip fractures will be dramatic as the population burden of fractures originates in men and women with osteopenia.

Men Women Eastern Mediterranean

Men Women Asia

Men Women Africa

Number of hip fractures in 1990 and those predicted in different regions of the world for 2025 and 2050

Joints
Cartilage consists of a single cell type, chondrocytes embedded in extracellular matrix made up of two major components: type II collagen which imparts tensile strength to the tissue and aggrecan that provides the ability of cartilage to resist compressive force. Orchestrated synthesis and turnover ensures and maintains the biochemical characteristic of the cartilage. Osteoarthritis is usually due to the During osteoarthritis, the most common form of arthritis, this regulation is disrupted by the expression of pro-inammatory molecules that provides the stimulus for the synthesis of matrix-degrading enzymes. These enzymes degrade aggrecan and collagen, resulting in the loss of cartilage and function. aging process, but osteoarthritis due to external factors has also been reported, for example in people whose activities involve joint stress, such as sports activities or even at work. Severe overweight can This chronic degeneration of the joints is already considered by the World Health Organization as one of the most disabling diseases in developed countries. For those who are affected, the also cause osteoarthritis. All joints can be affected, although it occurs more often on knee and hip joints and in the vertebral column. In advanced stages it is extremely painful. consequences are important because 80% have limitations in movement, and 25% can not perform their major daily activities of life.

160 140 120

Millions of people 2003 2008

Arthritis in the world According to worldwide estimation, arthritis affects 9.6% of men and 18% of women aged over 60 years.

100 80 60 40 20 0
India China US Brazil Japan

2013

Moreover, according to Datamonitor (2009), the arthritis prevalence has increased all over the world since 2003, such as in India (+2.3% per year) or Brazil (+2.1% per year), and will continue to grow, possibly even reaching 2.5% per year in India and 2.3% per year in Brazil.

Germany

France

Prevalence of osteoarthritis

Peptan

Collagen peptides for bone health

Collagen, which represents 90% of organic bone mass, has been identied as effective in the treatment of osteopenia. Several studies show that a daily intake of 10g of collagen peptides for 4 to 24 weeks increases bone mass density (1, 2).

this matrix, osteoblasts, the cells responsible for bone formation, are preferentially stimulated, instead of osteoclasts, the cells involved in bone resorption, this triggers bone formation (4, 5, 6).

Studies conducted by Rousselot with Peptan at Researchers have produced several keys for understanding how collagen peptides work. They have also shown that the extra-cellular matrix in which cells grow is decisive in their differentiation. Studies have demonstrated that when collagen peptides metabolites are present in the Physiology and Ingestive Behavior Laboratory, INRA-AgroParis Tech (Paris, France) and presented below, conrm these scientic ndings and provide new information about the effect of Peptan on bone metabolism (7).

A culture of osteoblasts (star-shape cells) and osteoclasts (round black cells) with Peptan.

Rousselot in vitro results

Peptan induces the differentiation of cells into osteoblasts mg/mL/min/DC In mixed cultures of osteoblasts and osteoclasts, the levels of alkaline phosphatase (ALP), a marker of bone formation, are increased when Peptan is present in the media. Since the number of cells does not increase with Peptan compared to the control group, this means that Peptan induces the differentiation of cells into osteoblasts, rather than osteoclasts. 0 BSA Peptan P Peptan B Measures of Alkaline Phosphatase 2 1,5 1 0,5 2,5 1 mg/mL 0,5 mg/mL * * * * * *

Peptan F

Measures of alkaline phosphatase (ALP) activity in bone cell culture with Peptan B (bovine), Peptan P (porcine) and Peptan F (sh) for 14 days, compared to BSA (standard proteins).

Peptan reduces bone resorption Moreover, when the culture is performed on a system which allows measurement of the osteoclast-mediated bone resorption, we can see that when Peptan is present the resorption area is reduced compared to the control.
A Cell culture with BSA standard protein (A) or with Peptan (B): osteoclasts activity is measured by the digestion of a calcium phosphate lm (white spot). B

Peptan induces differentiation into osteoblast, at the same time reducing the differentiation into osteoclast.

Rousselot in vivo results

The in vivo part of this study conrms but also reveals some completely new ndings. In this part, ovariectomized mice were used to simulate low bone mass density: osteopenia. This surgery actually slows down the bone (g/cm2) mineral density increase during the growth of the concerned mice. The mice were fed with or without Peptan for 12 weeks and various data was collected. The Peptan group had a 4g daily diet containing 2.5% (0.1g) of Peptan. 0,025 0,02 0,015 0,01 0,005 0
Increase of Bone Mineral Density (BMD) of the mice, in the control group (Control), the ovariectomized group fed without Peptan (Ovx), and the ovariectomized group fed with Peptan (Ovx + Peptan) after 12 weeks. Groups with different letters indicate signicant difference (p<0.05).

Peptan restores bone mineral density

BMD increase

a b

Control

Ovx

Ovx+Peptan

First, in ovariectomized mice fed with Peptan for 12 weeks, measures show restoration of the bone mineral density value close to the level of the control group (which was not ovariectomized).

Peptan increases bone size and solidity

Measure of the cortical area Measure of ultimate strength 31 30 1,2 (mm2) ab 29 28 27 26 1,1 Control Ovx Ovx+Peptan 25 Control Ovx Ovx+Peptan 1,25 a

Moreover, growth of the cortical zone (external area of the bone) of the femur, and a signicant increase in bone size, was measured in ovariectomized mice fed with Peptan, compared to ovariectomized mice not fed with Peptan. Due to this restoration, the ultimate strength of the bones (strength required to lead to a break) was signicantly greater for ovariectomized mice fed with Peptan.

ab

1,15

Finally, conrming the in vitro studies, in vivo studies have demonstrated that the bone resorption of ovariectomized mice fed Peptan, measured in terms of CTX production, is similar to the control group.

Measure of the cortical area and the ultimate strength, in the control group (Control), the ovariectomized group fed without Peptan (Ovx), and the ovariectomized group fed with Peptan (Ovx + Peptan).Groups with different letters indicate signicant difference (p<0.05) - ab is not signicantly different from a or b.

Peptan decreases bone resorption

Measure of CTX concentration 15

Peptan has been shown to be effective on bone metabolism by inducing differentiation (g/mL) and maturation of osteoblast and stimulating their activity. Bone turnover is thus modulated, preventing bone resorption during the natural phenomenon of bone loss (osteopenia) and increasing bone solidity. 0 a 10 b b

Control

Ovx

Ovx+Peptan

Measure of the Carboxy-Terminal collagen (CTX) peptides concentration, peptides fragments generated by collagen degradation, an important biochemical marker of bone resorption, in the control group (Control), the ovariectomized group fed without Peptan (Ovx), and the ovariectomized group fed with Peptan (Ovx + Peptan). Groups with different letters indicate signicant difference (p<0.05).

Peptan

A bioactive ingredient that reduces joint pain

Collagen peptides offer proven efcacy The benecial effects of collagen peptides have been reported in numerous research ndings. subjects with primary osteoarthritis of the knee were given 10g of collagen peptides daily. The results showed a signicant improvement in The clinical study conducted by Moskowitz (8), performed on various populations for a period ranging from 30 to 90 days, demonstrated a positive effect on pain reduction with a dose of 10g of collagen peptides per day. The effect was even more pronounced in patients suffering more severe symptoms. The effectiveness of collagen peptides has been also proven in athletes with activity-related joint pain. In a 24-week study conducted in 2008, 147 subjects who compete as part of a varsity team or Another more recent study assessed the efciency of collagen peptides in a randomized, double blind, controlled multicentre trial, in which 250 club sport were recruited and randomly assigned to two groups: one group received a liquid with 10g of collagen peptides, and the other received a placebo. At the end of the study, an assessment of participants by physicians showed statistically signicant changes with the dietary supplement collagen peptides: joint pain at rest, when walking, standing or carrying objects was reduced (10). knee joint comfort, and again subjects with the greatest joint deterioration beneted most (9).

The action mechanism of collagen peptides

Collagen peptides in cartilage Radioactivity (dmp/100 mg cartilage) 300 250 200 150 100 50 0 0 12
14

* *

Collagen peptides are accumulated in cartilage

Some explanations of this positive effect were revealed in the study by Oesser (11). He showed that more than 95% of 14C labeled collagen peptides taken orally is absorbed. However, he also showed that an important part is accumulated in cartilage of mice as early as 12 hours after ingestion. The radioactivity level was more than twice as high in cartilage following 14C labeled collagen peptide administration compared with the control group (14C labeled proline + collagen peptides).

24

36 48 60 Time (hours)

72

84

96
Time course of radioactivity in cartilage of mice subsequent to absorption of 14C labeled collagen peptides and 14C labeled proline in the control group. The mice received 10mg collagen peptides/g body weight. *p<0.05 between collagen peptides and control (11).

C labeled collagen peptides

Control

Type ll collagen secretion by chondrocytes in culture Type ll collagen (g/106 chondrocytes)

Collagen peptides induce the production of type II collagen by chondrocytes

In another study (12), the same author explains that the chondrocytes (cells of the cartilage) are sensitive to the products present in the extracellular matrix around them, and that the treatment of chondrocytes with collagen peptides in the medium leads to a dose-dependent production of collagen by those cells.
Time course of type II collagen secretion into supernatant of bovine chondrocytes cultured in basal medium or in medium supplemented with 0.5 mg/mL of collagen peptides. After 11 days of culture, type II collagen secretion was almost 2.5-fold higher in collagen peptides cultures. *p<0.01 between collagen peptides and control cultures (12).

1,8 1,6 1,4 1,2 1 0,8 0,6 0,4 0,2 0 6 8 Culture time (days) 11 * * *

Basal medium (control)

Medium containing collagen peptides

Conclusion
These different studies demonstrate the benets of collagen peptides on joint and bone health. The results indicate that collagen peptides could be useful in the prevention and treatment of osteopenia and osteoarthritis.

Regarding bones, Peptan has been shown to: Restore bone mineral density.  Increase the bones size, making them less brittle.  Stimulate osteoblast activity in spite of osteoclasts activity; the growth of the bone is stimulated.

Regarding joints, collagen peptides have been shown to:  Signicantly reduce joint pain, and subjects with the greater deterioration benet most.  Orally taken collagen peptides are accumulated in cartilage as soon as 12 hours after the ingestion and the cells of the joint, the chondrocytes, are responding to these extern collagen by producing a dose-dependant production of intern collagen.

Peptan
a completely characterized and scientically objectivised bioactive ingredient that helps to:

Prevent osteopenia Maintain bone health Prevent loss of bone solidity

Prevent osteoarthritis Reduce joint pain Prevent joint matrix degeneration

References
1.  Nomura, Y., Oohashi, K., Watanabe, M. and Kasugai, S. 2005. Increase in bone mineral density through oral administration of shark gelatine to ovariectomized rats. Nutrition, 21: 1120-1126. 2.  Wu, J., Fujioka, M., Sugimoto, K., Mu, G. and Ishimi, Y. 2004. Assessment of effectiveness of oral administration of collagen peptide on bone metabolism in growing and mature rats. Journal of bone and mineral metabolism, 22: 547-553. 3.  Karaguzel, G., Holick, M. 2010. Diagnosis and treatment of osteopenia. Rev Endocr Metab Disord, 11: 237-251. 4.  Mizuno, M. and Kuboki, Y. 2001. Osteoblast-related gene expression of bone marrow cells during the osteoblastic differentiation induced by type I collagen. Journal of biochemistry, 129: 133-138. 5.  Andrianarivo, A.G., Robinson, J.A., Mann, K.G. and Tracy R.P. 1992. Growth on type I collagen promotes expression of the osteoblastic phenotype in human osteosarcoma MG-63 cells. Journal of cellular physiology, 153: 256-265. 6.  Lynch, M.P., Stein, J.L., Stein, G.S. and Lian, J.B. 1995. The inuence of type I collagen on the development and maintenance of the osteoblast phenotype in primary and passaged rat calvarial osteoblasts: modication of expression of genes supporting cell growth, adhesion, and extracellular matrix mineralization. Experimental cell research, 216: 35-45. 7.  Guillerminet, F., Beaupied, H., Fabien-Soul, V., Tom, D., Benhamou, C-L., Blachier, F., Roux, C. and Blais, A. 2010. collagen peptides improves bone metabolism and biomechanical parameters in ovariectomized mice: an in vitro and in vivo study. Bone. 8. Moskowitz, R. 2000. Role of collagen hydrolysate in bone and joint disease. Seminars in arthritis and rheumatism, 30 (2): 87-99. 9.  Ruiz-Benito, P., Camacho-Zambrano, M.M., Carrillo-Arcentales, J.N., Mestanza-Peralta, M.A., Vallejo-Flores, C.A., Vargas-Lopez, S.V., Villacis-Tamayo, R.A. and Zurita-Gavilanes, L.A. 2009. A randomized controlled trial on the efcacy and safety of a food ingredient, collagen hydrolysate, for improving joint comfort. International journal of food sciences and nutrition, 12:1-15. 10.  Clark, K.L., Sebastianelli, W., Flechsenhar, K.R., Aukermann, D.F., Meza, F., Millard, R.L., Deitch, J.R., Sherbondy, P.S. and Albert, A.. 2008. 24-Week study on the use of collagen hydrolysate as a dietary supplement in athletes with activity-related joint pain. Current medical research and opinion, 24 (5): 1485-1496. 11.  Oesser, S., Adam, M., Babel, W. and Seifert, J. 1999. Oral administration of 14C labelled gelatine hydrolysate leads to an accumulation of radioactivity in cartilage of mice (C57/BL). Journal of nutrition, 129: 1891-1895. 12. Oesser, S. and Seifert, J. 2003. Stimulation of type II collagen biosynthesis and secretion in bovine chondrocytes cultured with degraded collagen. Cell tissue research, 311: 393-399. 13.  Pasco, J.A, Seeman, E., Henry M.J, Merriman, E.N, Nicholson, G.C, Kotowicz, M.A. 2006. The population burden of fractures originates in women with osteopenia, not osteoporosis. Osteoporosis Int, 17: 1404-1409.

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