Claude Laurent

Cross-linked hyaluronan used as augmentation substance for treatment of glottal insufficiency: Safety aspects and vocal fold function

Objective To examine safety aspects and vocal fold function after vocal fold augmentation with a cross‐linked hyaluronan derivative (hylan B gel) as compared with bovine collagen.

Hyaluronan and Healing of Tympanic Membrane Perforations. An experimental study

Hyaluronan, an extracellular high molecular weight polysaccharide, was applied onto experimentally performed tympanic membrane (TM) perforations in the rat. The hyaluronan-treated perforations closed faster than the untreated controls and showed less scar tissue. Initially the perforation became covered by a sheet of keratin and hyaluronan, which appeared to relax and orientate the retracted collagen fibres at the margin of the perforation. The hypertrophic stratified keratinizing squamous epithelium advanced within the hyaluronan-keratin cover and closed the perforation ahead of an approaching connective tissue. Three months after the closure, the hyaluronan-treated TMs appeared transparent in comparison to the opaque untreated controls. In histological sections the hyaluronan-treated TM was comparably thin and an organized collagen fibre layer was discernible. Hyaluronan of various concentrations (0.75, 1.0, 1.75 and 1.9%) and molecular weights (4 x 10(6) and 6 x 10(6)) were tested. The healing rate and degree of scar tissue was dependent on the concentration, but not on the molecular weight of the hyaluronan. Hyaluronan, at the higher concentration, healed the perforations faster and produced a less opaque TM than hyaluronan at low concentration. The number of applications of hyaluronan also influenced the healing rate; fewer applications resulted in a reduced rate. However, the quality of the healed TM was influenced to a lesser extent by the number of hyaluronan applications. Even single applications caused less scar tissue in comparison with untreated TMs. It is inferred from the study that hyaluronan, exogenously applied, might be valuable also in the clinical situation to enhance the healing of different types of TM perforation.

Cross-linked Hyaluronan (Hylan B Gel): a New Injectable Remedy for Treatment of Vocal Fold Insufficiency - an Animal Study

After injection of hylan B gel (Hylaform), the vocal folds of rabbits were studied by light microscopy at various time-points up to 12 months. Hylan B gel is a viscoelastic cross-linked and insoluble hyaluronan derivative which behaves as a soft gel. It is non-immunogenic, non-toxic and non-inflammatory and has already been used in plastic surgery for intradermal implantations. Macroscopic examination of the injected vocal folds revealed a bulging in six of the animals, but on microscopical examination the hylan B gel was encountered in all of them. Already after 1 month, and increasingly so up to 12 months, there was an ingrowth of new connective tissue, with collagen, fibroblasts and hyaluronan within the gel. No inflammatory reaction or any sign of a foreign body reaction was observed throughout the study. Hylaform may well become useful as an injection material for augmentation of vocal folds in the treatment of vocal fold insufficiency.

Cross-linked hyaluronan versus collagen for injection treatment of glottal insufficiency : 2-year follow-up.

Objectives—To evaluate the long-term (24 months) clinical performance (vocal fold function) and safety of hylan B gel as compared with bovine cross-linked collagen in the treatment of patients with glottal insufficiency. Material and Methods—In a prospective trial, 70 patients with glottal insufficiency due to unilateral vocal fold paresis (n=35) or atrophy (n=35) were randomized to received either hylan B gel (n=47) or collagen (n=23) injections into 1 vocal fold. Forty-two of the patients were examined 24 months after treatment. Evaluations were made based on patients’ subjective ratings, digitized videostroboscopic measurements, maximum phonation time and phonation quotient. Results—The patients’ self-ratings were significantly improved in both the hylan B gel and collagen groups. Videostroboscopic measurements of glottal closure were significantly improved for both groups. The hylan B gel group showed a trend towards less resorption at the injected vocal fold edge in comparison with the collagen group (p=0.05). No serious adverse events were observed. Twenty-eight patients dropped out of the study after 12 months: 18 had been re-injected or operated on with medialization laryngoplasty due to insufficient voice and 10 had either died of causes unrelated to the study or refused to attend follow-up. Conclusions—No long-term side-effects were found for either the hylan B gel or collagen groups after injection treatment. Both treatments resulted in significantly improved voice as rated by the patients and significantly improved glottal closure. Some resorption was noted for both substances, and ≈25% of the patients chose re-treatment 2 years after the initial treatment.

Hyaluronic acid reduces connective tissue formation in middle ears filled with absorbable gelatin sponge: an experimental study

Absorbable gelatin sponge, commonly used in otosurgery, was mixed with high-molecular-weight, highly concentrated hyaluronic acid. The mixture was introduced into the middle ear cavities of rats. The postoperative changes were evaluated by morphologic techniques after two months and three months. The middle ear cavity contained a loose mass of connective tissue with few cells, forming sail-like bridles between air-filled spaces. The filling material was easily detached from the surroundings. The structure of the middle ear mucosa and the tympanic membrane was well preserved. Compared with the findings in a previous study, in which absorbable gelatin sponge alone was used, the combination of gelatin sponge and hyaluronic acid caused significantly less structural alterations in the middle ear cavity. It is concluded that hyaluronic acid, a natural component in most tissues, markedly reduces the formation of the fibrous connective tissue known to develop in middle ear cavities filled with absorbable gelatin sponge alone.

Viscoelasticity of rabbit vocal folds after injection augmentation.

Objectives/Hypothesis Vocal fold function is related to the viscoelasticity of the vocal fold tissue. Augmentation substances used for injection treatment of voice insufficiency may alter the viscoelastic properties of vocal folds and their vibratory capacity. The objective was to compare the mechanical properties (viscoelasticity) of various injectable substances and the viscoelasticity of rabbit vocal folds, 6 months after injection with one of these substances.

Localization of hyaluronan in various muscular tissues

SummaryThe histochemical distribution of hyaluronan (hyaluronic acid, HYA) was analysed in various types of muscles in the rat by use of a hyaluronan-binding protein (HABP) and the avidin-biotin/peroxidase complex staining procedure. Microwave-aided fixation was used to retain the extracellular location of the glycosaminoglycan. In skeletal muscles, HYA was detected in the connective tissue sheath surrounding the muscles (epimysium), in the septa subdividing the muscle fibre bundles (perimysium) and in the connective tissue surrounding each muscle fibre (endomysium). HYA was heterogeneously distributed in all striated muscles. In skeletal muscles with small fibre dimensions (e.g., the lateral rectus muscle of the eye and the middle ear muscles), HYA was predominantly accumulated around the individual muscle fibres. Perivascular and perineural connective tissue formations were distinctly HYA-positive. In cardiac muscles, HYA was randomly distributed around the branching and interconnecting muscle fibres. In comparison, smooth muscle tissue was devoid of HYA.

Dextranomeres in hyaluronan (DiHA): a promising substance in treating vocal cord insufficiency.

The morphology of the rabbit vocal cord was studied after injecting a mixture of dextranomere microspheres in sodium hyaluronan solution (DiHA), two substances that are nonimmunogenic and biocompatible. DiHA is already in clinical use as a space filler in pediatric urology (Deflux R). Specimens from injected vocal cords were investigated by light microscopy at different time intervals up to 6 months after injection. On macroscopic examination a bulge was seen in the injected vocal cord, and on microscopic examination new fibrous connective tissue developed. Throughout the study a weak inflammatory reaction was observed, but no foreign body reaction. Hyaluronan disappeared from the injected site within 1 week, but the dextranomeres remained and were recovered intact up to 6 months after injection. The dextranomeres recruited fibroblasts that generated new collagen, resulting in endogenous soft tissue augmentation. It is conceivable that DiHA may become a useful injection material for treatment of vocal cord insufficiency.

The Catabolic Fate of Hyaluronic Acid

Part of the hyaluronic acid (HA) synthesized in peripheral tissues enters the blood circulation through the lymph. It is rapidly taken up by the endothelial cells in the liver (half-life in blood is 2.5-5.5 minutes) and degraded. Pure primary cultures of liver endothelial cells were obtained by a newly developed technique and used to follow the metabolism of the polysaccharide on the cell surface. At 37 degrees C the HA is effectively endocytosed and degraded to acetate and lactate. A radioassay specific for HA and sensitive in the nanogram range has been developed to follow the concentration of HA in serum. The normal level in man is 10 to 100 micrograms/l. Elevated serum levels of HA are seen in liver cirrhosis, rheumatoid arthritis and scleroderma indicating that both an impaired catabolism in the liver and an increased synthesis in the peripheral tissues can modify the HA level.

Localization and quantity of hyaluronan in urogenital organs of male and female rats

The histochemical distribution of hyaluronan was analysed in various urogenital organs of male and female (non-pregnant and pregnant) rats by use of a hyaluronan-binding protein and avidin biotin/peroxidase staining. Microwave-aided fixation was used to preserve the extracellular location of hyaluronan. The concentrations of hyaluronan in the different tissues were measured with a highly sensitive radio-assay. Hyaluronan accumulated predominantly in the connective tissue around smooth muscle fibres and in the subepithelial lamina propria. Abundant hyaluronan also occurred in perivascular and perineural connective tissue. In the female urogenital organs, hyaluronan content was high in the vagina and urinary bladder, and highest in the vagina during pregnancy. In the uterus, the surface epithelium of the endometrium stained intensely. In the ovary, the zona pellucida of the oocyte and the theca interna cell layer of the follicles and the follicular fluid of mature follicles exhibited prominent staining. The corpus luteum was devoid of hyaluronan, whereas enlarged corpora lutea of pregnancy exhibited weak, patchy staining. In male urogenital organs, staining for hyaluronan was absent from the testis and epididymis, whereas the erectile connective tissue of the penis stained intensely. The hyaluronan concentrations were high in penile tissue and urinary bladder, while testis, epididymis and the ductus deferens contained only little hyaluronan.