The Sun reports that US-based experts now think the smirk was caused by an underactive thyroid gland.
Known as hypothyroidism it can cause swollen hands, thinning hair and a lump in the neck — all visible in the portrait.
According to new research this was the condition triggered the enigmatic facial expression of Leonardo da Vinci’s famous model.
It was spurred by not eating enough dairy products, seafood and meat — and her earlier pregnancy.
The smile may be due to disability caused by muscle weakness, US medics suggested.
Professor of Medicine Dr Mandeep Mehra, medical director of the Heart & Vascular Centre at Brigham and Women’s Hospital in Boston, said: “The enigma of the Mona Lisa can be resolved by a simple medical diagnosis of a hypothyroidism-related illness.
“In many ways, it is the allure of the imperfections of disease that give this masterpiece its mysterious reality and charm.”
Previous examinations of the 16th century masterpiece have suggested she was suffering high cholesterol, heart disease — or even syphilis.
The limited treatments available at the time would have meant she would have died much younger.
Instead the new analysis published in Mayo Clinic Proceedings reveals noblewoman Lisa Gherardini — widely believed to be the subject — displays classic signs of hypothyroidism.
Her hands look swollen and it appears she’s losing her hair. She also has yellowed skin and what seems to be a goitre — or lump — in the neck.
Hypothyroidism in which the thyroid gland does not produce enough hormones can cause all these symptoms.
The researchers said: “The painting suggests a yellowish discolouration of the skin, which is known to occur in hypothyroidism.
“The black veil that hangs below what appears to be a large forehead indicates a receding hair line, with hair that appears to be thinned.
“A complete lack of eyebrows or other hair throughout the pale skin further supports this diagnosis, and cascading hair down the side appears coarse in character.
“Curiously, a close look at the neck does insinuate the possible presence of a diffuse enlargement such as a goitre.”
They said swelling in the right hand further supports the theory as this is often noted in advanced stages of hypothyroidism.
The researchers said: “In this circumstance, if Lisa Gherardini was indeed suffering from severe hypothyroidism or its consequences, the mysterious smile may at one level be representative of some psychomotor retardation and muscle weakness leading to a less than fully blossomed smile.”
The condition can be caused by a lack of iodine — found in dairy products, seafood, meat, bread and eggs.
This was a common problem among Italians during the Renaissance. Resulting goitre were commonly depicted in paintings and sculptures.
The researchers said: “Thus, the diet was one that was often iodine deficient and more importantly, the eating habits promoted the development of goitre.”
Dr Mehra and Dr Campbell also pointed out Lisa Gherardini gave birth shortly before she began sitting for da Vinci.
Pregnancy can also contribute to inflaming the gland.