Study Aim

In vitro laboratory studies were undertaken by the Australian Centre for Complementary Medicine Education and Research (ACCMER), a joint venture of the University of Queensland and Southern Cross University. The aim of the studies was to compare selected biological activities of Healtheries' Biolane™ (Green Lipped Mussel Extract) versus the biological activities in a range of well known complementary anti-arthritic agents.

Sample Preparation

All samples were subjected to two enzyme digestions to simulate in vivo digestive processes. Various components of gastrointestinal secretions may have an impact on potential actives within a product. In this study we used a two stage in vitro model based on gastric and duodenal secretions. The digests prepared from the samples were a pepsin digest containing the prominent gastric enzyme pepsin and a complete digest where the sample was exposed to pepsin followed by pancreatic enzymes.

Assays
Assays assessed:

• cholesterol synthesis inhibition
• anti oxidant capacity

Inhibition of the following components of inflammatory pathways

• tissue necrosis factor alpha (a)
• Cox-2 expression
• prostaglandin E2 (PGE2)
• phospholipase A2 (PLA 2) - also associated with platelet aggregabillity
• platelet aggregation inhibitory activity
• fibrinolytic activity

Synopsis of Results

Comparison of aggregate in vitro data - Chondroitin Sulphate (CS) vs Glucosamine Sulphate (GS) vs Glucosamine sulphate:Chondroitin sulphate (GS/CS) vs Lyprinol™ vs Healtheries Biolane™ (Green Lipped Mussel Extract)

Test	Anti-arthritic agent
	CS	GS	GS/CS	LYP	GLME
Cholesterol biosynthesis inhibition	●	●	●	●	●
TNFa inhibition	●	●	●	○	●
Cox-2 inhibition	○	●	●	ND	●
PGE2 inhibition	○	○	●	●	●
PLA2 inhibition	●	●	○*	●	●
Oxygen radical absorbance capacity - antioxidant (ORAC)	○	○	○	○	●
Fibrinolytic activity	○	●	●	○	○
Anti-platelet aggregation activity	○	●	○	●	○
Note ● denotes activity present ○ denotes activity not found ○* denotes activity not found at low concentration but present at higher concentration ND = not done (test unable to be performed due to assay artefact) The results show that Healtheries Biolane™ (Green Lipped Mussel Extract) demonstrates the most comprehensive range of activities across the suite of in vitro tests performed when compared with the other agents that were tested.

Relevance of the in vitro Biolane green lipped mussel extract (GLME) findings to putative Osteoarthritis pathomechanisms

The vascular theory of osteoarthritis causation is based on epidemiological, laboratory, experimental and clinical findings that support the concept that compromised microcirculation in affected joints initiated through a combination of inflammation and imbalance between coagulation and fibrinolysis can initiate and perpetuate the disease. One implication of this theory is that in order to treat more than just painful symptoms ie to slow or halt the disease process will require a range of biochemical activities to effectively break the web of pathology. It has been proposed that these should include a number of key activities such as anti-inflammatory, anticoagulant, fibrinolytic and lipolytic activities. If these are realised then chondroprotection is likely to ensue.

The use of highly potent anti-inflammatory agents, particularly the Cox-2 inhibitors would appear to be at odds with the known data showing that patients with osteoarthritis are at greater risk of thrombotic episodes than those without the disease. Cox-2 inhibitors pose a theoretical risk of increased thrombotic complications in many patients with osteoarthritis who already have cardiovascular risk factors. The recent removal of Vioxx from the market and restrictions governing the use of Celebrex - the Cox-2 market leaders, based on these concerns would appear to support this proposition.

A successful anti-arthritic agent should have multiple low level activities that address a range of disease drivers while avoiding the serious side effects that frequently accompany massive disruption of major biochemical pathways such as total Cox-2 inhibition.

The current in vitro study has shown that GLME has a range of activities that the vascular theory of osteoarthritis causation predicts as desirable for disease treatment. These include anti-inflammatory activity through its inhibition of TNFa, PGE2, Cox-2 and PLA2 in addition to its anti-oxidant activity. It is not critical for these activities to be at extremely high levels. In fact it is advantageous from the perspective of a low side effect profile that they should not be so. It is the multiplicity of activities that is of greater importance. In addition to anti-inflammatory activity, GLME also has in vitro activities that can reduce thrombotic risk (decreased PLA2 and cholesterol biosynthesis inhibition activity in addition to decreased platelet aggregability - that is also assisted through cholesterol biosynthesis inhibition) and enhance the removal of blood clots (mild fibrinolytic activity).

The results obtained in this study indicate that GLME has in vitro activities in key areas associated with arthritis pathophysiology. In addition, across the range of in vitro tests performed in this comparative study, Healtheries Biolane™ green lipped mussel extract demonstrated a more extensive range of potential anti-arthritic activities in comparison to the other agents tested.

It is important to note that these results apply specifically to Biolane™ (Green Lipped Mussel Extract) and can not be extrapolated to include other green lipped mussel extracts.