The analysis of mitochondrial disease involves the integration of different medical specialties organized in subsequent stages. The results of laboratory and instrument analysis are integrated with clinical examination, family and natural history of the patient. The clinical evaluation of children with suspected mitochondrial pathology is carried out in the Metabolic Unit of the Division of Child Neurology at the National Neurological Institute, under the supervision of Dr. Graziella Uziel. The clinical evaluation of adult patients is carried out by Dr. Massimo Zeviani and Dr. Costanza Lamperti(for more details see OUR SERVICES). These diseases strike primarily the brain and skeletal muscle. Therefore neuroradiological evaluation based on brain Magnetic Resonance Imaging (MRI) and/or mass spectrometry, and neurophysiological studies including Evoked Potentials and EEG are routinely performed. However, the most fundamental exam in most cases is the muscle biopsy, which is used for morphological studies by light and electronic microscopy and provides the biological material indispensable for biochemical analysis of the respiratory chain and for mutation analysis on candidate genes. Muscle biopsy is particularly useful in pediatric cases, in which symptoms are often unspecific and it is therefore not possible to establish a sure analysis only on the basis of clinical examination.

Analysis on body fluids
Blood and urine tests are useful for a preliminary screening.

Metabolic screening for suspected mitochondrial disease
Assay Sample* Comments
Glucose B
Ions and electrolytes B
Lactata, Pyruvate, Alanine B, CSF Do not use tourniquet; intercurrent infections, exercise and stress can alter the results
Lactate/Pyruvate ratio B Useful to determine the point of the metabolic block
Ammonia B
Aminoacids B,U, CSF Urinary aminoacids can be collected during fast conditions or after a meal. Generalized aciduria can suggest a mitochondriopathy
Organic acids U Take a 24h urine collection. Keep urine a t 4°C in sterile conditions with no preservants
Carnitine and derivatives B Free, short-chain esters, long-chain esters, and total carnitine should be measured separately
Ketones B,U Should be measured during a metabolic crisis
B= blood U=urine CSF=cerebro-spinal fluid

Results of these analyses can already orient the specialist toward the correct diagnosis specialist. Negative results, however, do not completely exclude the hypothesis of a mitochondrial disease. If the suspect persists, it is advisable to repeat the assays under energy stress, for instance during a spontaneous metabolic crisis or by inducing a metabolic stress (by prolonged fasting , glucose or fatty acid load, etc). These tests must be carried out under close supervision in a specialized hospital.

Muscle Biopsy
This is now carried out with the use of a surgical needle, which is less painful and less invasive then the "open" surgical technique. Needle biopsy is carried out under local anesthesia in adult patients and under general anesthesia in children. Suture stitches are not necessary, the resulting scar is very small and leaves no lasting effect on the biopsied muscle. It is of utmost importance that the exam is carried out in a specialized institution, that can guarantee the correctness and safety of the technique, the proper conservation of the material and the completeness of the analysis.

The following investigations can be carried out on the muscular tissue from the biopsy:

In particular cases, electron microscope examination can visualize the mitochondrial internal structure, which may be abnormal in mitochondrial disorders.

Skin Biopsy
Cell culture of fibroblasts obtained from skin biopsy can be useful in many cases:

Our center possesses a unique patrimony in Italy of more than 1000 lines of fibroblasts from patients with congenital metabolic pathologies.

All of these exams are carried out at our institute.

Ragged red fibers are a typical hallmark of mitochondrial myopathies

Histochemical staining for succinate dehydrogenase: the “ragged blue fiber” is equivalent to a ragged-red fiber: increased staining indicates abnormal proliferation of mitochondria

Numerous muscle fibers stain negative (white) to the histochemical staining specific to cytochrome c oxidase

Ultrastructural abnormalities: electron microscopy examination shows mitochondria which are altered in shape, size and internal structure