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PubMed1999

Effect of hyperbaric oxygen and medicinal leeching on survival of axial skin flaps subjected to total venous occlusion.

This study evaluates the effect of hyperbaric oxygen and medicinal leeching on axial skin flaps subjected to total venous occlusion. Axial epigastric skin flaps (3 x 6 cm) were elevated on their vascular pedicles in 40 male Wistar rats. Total venous occlusion was achieved by division of all veins draining the skin flap. Arterial inflow was left intact. Animals were randomly assigned to one of five groups: sham (n = 8); control, total venous occlusion only (n = 8); occlusion with hyperbaric oxygen (n = 8); occlusion with leeching (n = 8); occlusion with leeching and hyperbaric oxygen (n = 8). The hyperbaric oxygen protocol consisted of 90-minute treatments, twice daily, with 100% O2 at 2.5 atmospheres absolute for 4 days. The leeching protocol consisted of placing medicinal leeches on the congested flaps for 15 minutes, once daily, for 4 days. Laser Doppler measurements of flap perfusion were recorded preoperatively, postoperatively, and on postoperative days 1 and 3. The percentage of flap necrosis was evaluated on postoperative day 3. Mean percentage necrosis and mean laser Doppler readings were compared between both groups. The flaps in the sham group demonstrated 99 percent survival, whereas the flaps in the occlusion-only group demonstrated 100 percent necrosis. The flaps in the occlusion with oxygen, the occlusion with leeching, and the occlusion with oxygen and leeching groups demonstrated 1, 25, and 67 percent survival, respectively. Sham laser Doppler readings remained within normal limits. Laser Doppler readings in the occlusion-only and the occlusion with oxygen groups decreased to negligible levels on postoperative day 1, and on postoperative day 3 no perfusion was demonstrated. In both the occlusion with leeching and the occlusion with leeching and oxygen groups, there was also a significant decrease in laser Doppler measurements after surgery, but perfusion remained stable throughout the remainder of the study. This study demonstrates that hyperbaric oxygen alone is not an effective treatment for skin flaps compromised by total venous occlusion. The combination of leeching and hyperbaric oxygen treatment of total venous occlusion results in a significant increase in flap survival above that found with leeching alone. It appears that hyperbaric oxygen is effective because of the venous outflow provided by leeching as demonstrated by laser Doppler flow readings.

Animals↗
PubMed1992

Aeromonas hydrophila infections following use of medicinal leeches in replantation and flap surgery.

Aeromonas hydrophila infections are a recognized complication of postoperative leech application, and can occur with measurable frequency in populations of patients treated with leeches. We review 11 previously reported leech-related Aeromonas infections and analyze seven unreported cases. These infections range from minor wound complications to extensive tissue loss and sepsis. Often, these infections followed leech application to tissue with questionable arterial perfusion. Onset of clinical infection in these patients ranged from within 24 hours of leech application to 10 days or more after leech application. Late infections may represent bacterial invasion from colonized necrotic tissue. Based on these observations, we recommend that leech applications be restricted to tissue with arterial perfusion to minimize contamination of necrotic tissue. We also recommend that patients treated with leeches receive antibiotics effective against Aeromonas hydrophila before leech application. Patients treated with leeches and discharged with eschars or open wounds might benefit from oral antibiotic therapy until wound closure. These precautions may minimize or eliminate this complication of leech use.

Adolescent↗
PubMed1998

The failing flap in facial plastic and reconstructive surgery: role of the medicinal leech.

OBJECTIVE: To review the use of the medicinal leech, Hirudo medicinalis, in salvaging the failing, venous-congested flap. A protocol for the use of leeches is presented. Four illustrative cases of failing flaps (pectoralis major, midline forehead, and temporalis) are presented. STUDY DESIGN: Literature review comprised of MEDLINE search 1965 to present. Retrospective review of four cases involving the management of the failing, venous-congested flap. METHODS: A retrospective review of four cases of failing, venous-congested flaps was performed. RESULTS: The authors' experience, as well as the data from the reviewed medical literature, demonstrates the importance of early intervention in order to salvage the failing, venous-congested flap. Leeches are an immediate and efficacious treatment option. CONCLUSIONS: 1. Review of the literature indicates that the survival of the compromised, venous-congested flap is improved by early intervention with the medicinal leech. H medicinalis injects salivary components that inhibit both platelet aggregation and the coagulation cascade. The flap is decongested initially as the leech extracts blood and is further decongested as the bite wound oozes after the leech detaches. 2. When a flap begins to fail, salvage of that flap demands early recognition of reversible processes, such as venous congestion. The surgeon must be familiar with the use of leeches and should consider their use early, since flaps demonstrate significantly decreased survival after 3 hours if venous congestion is not relieved. In the four cases presented, a standardized protocol facilitated early leech use and provided for the psychological preparation of the patient, availability of leeches, and an antibiotic prophylaxis regimen. 3. The complications associated with leech use can be minimized with antibiotic therapy, wound care, and hematocrit monitoring. 4. The use of the medicinal leech for salvage of the venous-congested flap is a safe, efficacious, economical, and well-tolerated intervention.

Aged↗
PubMed2001

Cellular localization of a renin-like enzyme in leeches.

OBJECTIVES: The purpose of this study was to localize in leeches the renin-like enzyme previously characterized as well as the leech angiotensin-converting like enzyme (ACE). METHODS: Immunocytochemical as well as whole mount experiments were performed with an antibody raised against a fragment of the leech renin-like enzyme (VLWAAEKTQLDTGSS) and with anti-leech ACE. RESULTS: Anti-leech renin stains the vascular pole of the glomerulus and the afferent arteriole of the rat kidney. Immunostaining of leech sections revealed labeling in neurons and glial cells of the central nervous system (CNS), immunocytes and the nephridial canal, canaliculi and the periphery of the ciliated funnel, as well as the epithelium lining nephridia. Co-localization between antibodies raised against this fragment and a fragment of leech angiotensin-converting enzyme was demonstrated in neurons and glial cells of the leech CNS, as in vertebrates MAIN FINDING: Leech renin is localized in leeches like in vertebrate in the excretory system and in the nervous system. CONCLUSIONS: Our findings suggest the presence of a renin-angiotensin system involved in osmoregulation in leeches.

Amino Acid Sequence↗
PubMed2001

[Preliminary studies on process and ultramicro-pulverization on pharmacodynamics of Leech].

OBJECTIVE: To compare pharmacodynamics of decoction and powder of Leech; To identify effect of pharmacodynamics of Leech with new methods of process and ultrmicro-pulverization. METHOD: The size of ultramicor-powder of Leech and new method processed Leech is determined by BT-1500 size distributive instrument. It is compared that the pharmacodynamics of Leech decoction and powder as well as ultramicro-powder with new method processed of 1/2 decoction dose by anticoagulation and antithrombus action of mice. The safety of different samples is identified. RESULT: The pharmacodynamics of samples is as ultramicro-powder of Leech with new method processed > ultrmicro-powder of Leech > powder of Leech > decoction of Leech. CONCLUSION: The ultramicro-powder of Leech with new method processed has better pharmacodynamic effect and the smell of Leech is improved.

Animals↗
PubMed2002

Evaluation of performance characteristics of the medicinal leech (Hirudo medicinalis) for the treatment of venous congestion.

Medicinal leeches (Hirudo medicinalis) are a standard treatment for venous congestion, a complication that can occur after reconstructive surgery. If the cause of venous congestion cannot be surgically corrected, then medicinal leeches are used to temporarily increase perfusion levels and maintain physiologic requirements within the congested tissue. Leeches increase perfusion within congested tissue by actively drawing off blood as a bloodmeal. Furthermore, the leech bite continues to bleed and relieve congestion after detachment because of the anticoagulation effects of leech saliva left behind in the bite. In a porcine model, a 10 x 10 cm cutaneous flank flap was congested by clamping the venae comitantes. Four medicinal leeches were allowed to attach to the congested flap, and parameters of active feeding and passive bleeding after detachment were recorded. The average bloodmeal volume for the medicinal leeches was 2.45 ml. Average passive bleeding for the first 2 and 4 hours after leech detachment totaled 2.21 and 2.50 ml, respectively, with 90 percent of passive bleeding occurring within 5 hours after detachment. Laser Doppler imaging indicated that the spatial arrangement of surface perfusion increases were localized to a 1.6-cm-diameter circle around the leech head (bite) and corresponded well with the visual return of normal skin tones to the same area. This study provides a realistic and quantitative estimate of the spatial and volumetric characteristics of leech feeding and passive bleeding using a clinically relevant model of acute, severe congestion.

Animals↗
PubMed1991

Serotonin in the leech central nervous system: anatomical correlates and behavioral effects.

1. Serotonin is sequestered by a limited population of identified neurons in the 32 ganglia of the leech nervous system. A major fraction of the serotonin in each ganglion is contained in the paired Retzius cells, colossal effector neurons whose size varies longitudinally. The 5 other classes of identified serotonin-containing neurons, one effector cell and 4 interneurons, are approximately twice as numerous in anterior as in posterior ganglia. 2. We dissected 6 longitudinal samples from the ventral nerve cords of hungry Hirudo medicinalis, and measured their serotonin content using high pressure liquid chromatography with electrochemical detection. A consistent neurochemical pattern emerged in which segmental ganglia 2-4 had the highest quantity of serotonin: 18.51 pmol per ganglion. The anterior cerebral ganglion contained 14.78 pmol, and the content of the 4 posterior samples, segmental ganglia 7-10, 12-15, 17-20 and the caudal ganglion, decreased continuously from 16.35, 15.08, 10.75 to 2.51 pmol per ganglion, respectively. Morphometric analyses indicated that this pattern of ganglionic serotonin correlated primarily with longitudinal variations in the number of serotonin neurons per ganglion and secondarily with volume of the Retzius cells. Retzius cell volume correlated highly with the mass of their innervated body segments both of which are largest in mid-body domains. 3. Serotonin expresses leech feeding, and its ganglionic levels are a potentially useful index of behavioral state. We measured serotonin in the ganglionic samples from hungry and satiated leeches. The samples from recently fed animals contained 28% less serotonin than those from hungry ones. The amounts of serotonin in the cerebral and all the segmental samples from satiated leeches were significantly lower than equivalent samples of hungry animals. A similar pattern of depletion was seen in leeches which fed for a prolonged period (90 to 120 min) rather than the normal period of 30 min. 4. The effects of ingestion on serotonin-containing neurons was examined with the glyoxylic acid-induced histochemical fluorescence. The levels of fluorescence in all serotonin neurons in fed leeches were consistently lower than those in equivalent neurons in hungry animals, corroborating the ganglionic decrease in serotonin in satiated leeches. 5. To examine effects of body wall distension on serotonin levels, hungry leeches were fed to satiation, and half of them were relaxed by removing their distending blood meals. After 6 weeks, ganglionic serotonin in leeches with relaxed bodies was 21% higher than in those with distended bodies. 6. Ingestive behavior depletes serotonin from leech neurons and body wall distension appears to interfere with its synthesis.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

Leeches: controlled trial in venous compromised rat epigastric flaps.

Leeches were studied for their efficacy to improve survival of venous compromised rat skin flaps. In 22 rats, bilateral epigastric island flaps were created and subjected to 6 h of venous occlusion. One flap in each animal was randomised to leech treatment, while the contralateral flap served as its own control. Flap survival, leech feeding time, weight gained by the leech and bleeding time from leech bites were measured. The area of flap survival was significantly increased in leech treated flaps compared to contralateral controls (n = 22 pairs, p = 0.03; Wilcoxon signed rank test). Weight gained in the feeding leech averaged 1.3 +/- 0.2 g (n = 18). Leech feeding time was 107 +/- 13 min (n = 18). Bleeding time from each leech bite averaged 79 +/- 12 min (n = 18). Hence, the extent of flap necrosis resulting from venous impairment can be partly diminished by leech treatment until definitive surgical venous revascularisation.

Animals↗
PubMed1991

Leech therapy: when once is not enough.

Venous congestion is a constant threat in the survival of free flaps, pedicled flaps, tissue replantations and traumatized tissue. Leech therapy has proved effective in salvaging much of these compromised tissues. To be effective, leeches must be both readily available and hungry, requirements not always easily met. Our investigation seeks to establish a means to return sated leeches to their previous unfed, hungry state for reuse. Sated leeches were purged of their blood meals by placement in 3% hypertonic saline or by gentle finger pressure, then exposed to serotonin 0 microM (control), 10 microM, 30 microM or 90 microM for 20 min. Subsequent rebiting/refeeding was observed and analysed. Leeches bathed in serotonin rebit or reattached at nearly four times the rate of unexposed leeches; 30% vs 8% respectively. Biting, however, is not refeeding. Overall, 41 leeches were treated with serotonin with four (10%) refeeding. Those that refed consumed a significantly smaller blood meal than the initial feeding; 50% +/- 47% SD vs 348% +/- 143% SD of original body weight. None of the control group refed. As a method for routinely reusing leeches, serotonin bathing cannot be recommended. In the immediate postoperative period with the sudden emergence of venous congestion requiring leech therapy, but with an inadequate number available, this 20% [corrected] refeed rate after 10 microM serotonin exposure could potentially determine the success or failure of the flap/replantation until fresh leeches are made available.

Animals↗
PubMed2003

Water and sediment toxicity assessment by use of behavioural responses of medicinal leeches.

Behaviour has been shown to be a sensitive indicator of chemically induced stress and pathology in aquatic organisms. However, ecotoxicological investigations on medicinal leech are restricted because of scarce leech resources in natural waters. We used artificially bred medicinal leech (Hirudo medicinalis L.) of two different ages: young (1-2 weeks old) and adult leeches (1 year old). Animals were exposed to: (1) waters of Lake Drukshiai-the cooler of Ignalina Nuclear Power Plant, (2) sediments of the river Nemunas and (3) solution of heavy metal model mixture (HMMM). The following behavioural responses of leech were investigated: mobility (number of moving individuals within certain periods of time), avoidance response (number of individuals escaping the tested water or sediments) changes in body shape (contractions of some muscles, abnormal position of suckers) and feeding activity (longevity of attachment process, interruptions of feeding bouts, size of blood meal). Mobility of young leeches was increased in the tested waters of Lake Drukshiai, whereas adult leeches showed no changes of this index. Avoidance response as well as impaired feeding activity (prolonged attachment process or completely suppressed attachment reflex, decreased size of blood meal) was observed in young leeches exposed to the waters from Lake Drukshiai and in adult animals exposed to all three kind of trials mentioned above. Avoidance response and changes in mobility were recorded during the first hour of exposure to tested samples; therefore, these responses can be used as an express method for water and sediment pollution assessment. Impaired feeding activity was recorded after 1-3-week exposure: this response may be used in assessing a chronic toxicity of pollution. Medicinal leech, due to its sensitivity, simplicity of measured indices and ease of laboratory maintenance, can be used as a test organism in ecotoxicity studies.

Animals↗
PubMed1999

Aeromonas species isolated from medicinal leeches.

Aeromonas hydrophila infections are a recognized complication of the use of medicinal leeches. The authors performed an experiment designed to find a safe and practical way to sterilize the leech gut of pathogenic organisms. Leeches were incubated for a 12-hour period in solutions of antibiotic effective against A. hydrophila. The incubations in the antibiotic solutions failed to eradicate pathogenic bacteria from the gut of the leeches. The authors examined cultures of bacteria isolated from the guts of the commonly used Hirudo medicinalis (European leech) and found a wide variety of pathogenic organisms. A. hydrophila is widely believed to be the most common enteric pathogen, but the authors found A. sobria more frequently in their experiment. They also cultured the guts of the leech H. michaelseni recently used clinically in South Africa. A. caviae was the most common pathogen encountered in these leeches. A. caviae and A. sobria cause a spectra of disease similar to A. hydrophila. The authors endorse the current recommendation that all patients who have leech therapy for congested flaps or replants receive broad-spectrum prophylactic antibiotics. This appears to be the safest and simplest way to prevent leech-related infections.

Aeromonas↗
PubMed1993

The complete amino acid sequence of a hirudin variant from the leech Hirudinaria manillensis.

Unlike the European leech Hirudo medicinalis, the Asian jawed leech Hirudinaria manillensis is specialized for feeding on mammalian blood. In the salivary glands of both these leeches, there is a potent inhibitor of thrombin, called hirudin, which acts as an anticoagulant. We have reported previously the isolation and purification of a variant of hirudin, called bufrudin, from the head portions of Hirudinaria. In the present study, the complete amino acid sequence of bufrudin was determined by automated Edman degradation of peptide fragments generated after cleavage of protein with trypsin or thermolysin. Comparison of the primary structure of bufrudin, with hirudin HV1, show about 70% sequence identity with deletion of two amino acids, but the key amino acids at the C-terminus, involved in the inhibition of thrombin, are conserved. However, similar sequence comparison of bufrudin with hirullin P18, a hirudin variant isolated from the same leech species but from whole leech, instead of heads, reveals even less sequence identity of about 60%. From the amino acid sequence, it is suggested that the conformation of the C-terminal portion of bufrudin may be significantly different from hirullin P18, but similar to hirudin HV1, upon its interaction with thrombin. These results indicate that, as with Hirudo leech, various isoforms of hirudin also exist in Hirudinaria leech, with a significant change occurring in the structure of the molecule during the evolution of leeches.

Amino Acid Sequence↗
PubMed1992

The use of leeches for treatment of venous congestion of the nipple following breast surgery.

The medicinal leech, Hirudo medicinalis, played a central role in the evolution of medieval and folk medicine. Today, for the first time in history, the leech actually has a real and valuable purpose in medicine as a useful adjunct for the plastic surgeon: It provides relief of venous congestion. For over 2000 years, leeches were needlessly applied for a multitude of maladies as an adjunct to blood-letting. Their use in Europe peaked between 1830 and 1850, however, shortages led to a subsequent decline in leech application. Today there is a real clinical use for leeches which had led to a resurgence in their use in plastic surgery. Plastic surgeons use leeches in microsurgery to salvage congested flaps, whose viability is uncertain due to venous congestion. We present our experience with two patients where leeches were used to treat isolated venous engorgement of the nipple following breast surgery. Leech therapy is painless, well tolerated, and does not result in significant scarring. Prompt initiation of treatment is mandatory and produces dramatic resolution of venous congestion.

Adult↗
PubMed1994

Experiments on the possible role of leeches as vectors of animal and human pathogens: a light and electron microscopy study.

The presence and survival of pathogens inside the gut of leeches were studied by means of light and electron microscopy. In African leeches from Cameroon, blood was serologically positive for human immunodeficiency virus (HIV) and hepatitis B; blood of Hirudo medicinalis bought in German pharmacies contained up to 11 different species of bacteria. In experiments done at low (3 degrees C) and high (22 degrees, 32 degrees C) temperatures, it was shown that ingested red and white blood cells survive for long periods. The time was prolonged to at least 6 months in cases in which the leeches were stored at 3 degrees C. The same effect occurred with pathogens. Bacteriophages (viruses of bacteria) and bacteria persisted in large numbers for at least 6 months in the gut of experimentally infected leeches. Protozoan parasites such as Toxoplasma gondii, Trypanosoma brucei brucei, or Plasmodium berghei were even capable of reproducing inside the gut of the leech. In the case of Plasmodium parasites, this proceeded at low (3 degrees C) and high (22 degrees C) temperatures until all erythrocytes were used up. These parasites survived as long as the erythrocytes and lymphocytes were of good shape, i.e., around 5-6 weeks p.i. Single stages survived longer, especially at low temperatures. However, electron microscopy studies gave no hint of penetration of such pathogens into the unicellular salivary glands, which would initiate a direct transmission. Such transmission, however, is possible--many fish leeches directly transmit several blood parasites--when the leeches are squeezed during skin attachment or when they are manipulated by dropping salt solution on their backs while they are sucking. Consequently, the leech is a potential vector of many pathogens, especially in regions with an endemic spread of human and/or animal pathogens.

Animals↗
PubMed1986

Severe anaemia and death due to the pharyngeal leech Myxobdella africana.

Six cases of pharyngeal leech (identified as Myxobdella africana) are reported from a highland area of North Kenya. Three patients were severely anaemic and one died. There have been sporadic reports of leech infestation of man in East and Central Africa but these reports lack correct identification of the leech. This paper includes a detailed account of the systematics of the leech and of its habitat. The leeches were ingested accidentally by drinking water from two infested water sources. The major symptoms were a feeling of something in the throat, epistaxis and haemoptysis. On examination, pharyngeal blood was a consistent feature although the leech was rarely seen on initial inspection. Examination under anaesthesia was usually successful in locating and removing the leech. Prompt blood transfusion was life-saving in two cases. The water sources were used by both cattle and man; the cattle may also be parasitized by leeches. Methods of improving the water sources were discussed with community leaders.

Adolescent↗
PubMed1993

Bacteriological investigation of the occurrence and antibiotic sensitivities of the gut-flora of the potential southern African medicinal leech, Asiaticobdella buntonensis (Hirudinidae).

The lack of availability of medicinal leeches is a major impediment to the widespread use of leech therapy for treatment of congested flaps and replants in southern Africa. An investigation into the suitability of an alternative leech, the indigenous southern African leech, Asiaticobdella buntonensis, was therefore started. The risk of hospital-acquired infection related to the use of leeches and the antibiotic sensitivities of bacteria isolated from the gastro-intestinal tract of wild-caught leeches were investigated. Eleven bacterial genera were isolated but Aeromonas were most frequently isolated, occurring in 82% of microbiological samples. All were sensitive to cefotaxime and amikacin. The gut-flora and their sensitivities to 19 antibiotics were similar to those reported for the traditional medicinal leech, Hirudo medicinalis. These results emphasize the need to anticipate unusual infections when prescribing prophylactic or curative antibiotics in the course of leech therapy.

Aeromonas↗
PubMed2001

Evidence for an endocannabinoid system in the central nervous system of the leech Hirudo medicinalis.

In invertebrates, like Hydra and sea urchins, evidence for a functional cannabinoid system was described. The partial characterization of a putative CB1 cannabinoid receptor in the leech Hirudo medicinalis led us to investigate the presence of a complete endogenous cannabinoid system in this organism. By using gas chromatography-mass spectrometry, we demonstrate the presence of the endocannabinoids anandamide (N-arachidonoylethanolamine, 21.5+/-0.7 pmol/g) and 2-arachidonoyl-glycerol (147.4+/-42.7 pmol/g), and of the biosynthetic precursor of anandamide, N-arachidonylphosphatidyl-ethanolamine (16.5+/-3.3 pmol/g), in the leech central nervous system (CNS). Anandamide-related molecules such as N-palmitoylethanolamine (32.4+/-1.6 pmol/g) and N-linolenoylethanolamine (5.8 pmol/g) were also detected. We also found an anandamide amidase activity in the leech CNS cytosolic fraction with a maximal activity at pH 7 and little sensitivity to typical fatty acid amide hydrolase (FAAH) inhibitors. Using an antiserum directed against the amidase signature sequence, we focused on the identification and the localization of the leech amidase. Firstly, leech nervous system protein extract was subjected to Western blot analysis, which showed three immunoreactive bands at ca. approximately 42, approximately 46 and approximately 66 kDa. The former and latter bands were very faint and were also detected in whole homogenates from the coelenterate Hydra vulgaris, where the presence of CB1-like receptors, endocannabinoids and a FAAH-like activity was reported previously. Secondly, amidase immunocytochemical detection revealed numerous immunoreactive neurons in the CNS of three species of leeches. In addition, we observed that leech amidase-like immunoreactivity matches to a certain extent with CB1-like immunoreactivity. Finally, we also found that stimulation by anandamide of this receptor leads, as in mammals, to inhibition of cAMP formation, although this effect appeared to be occurring through the previously described anandamide-induced and CB1-mediated activation of nitric oxide release. Taken together, these results suggest the existence of a complete and functional cannabinoid system in leeches.

Adenylyl Cyclases↗
PubMed1996

Exploring the use of the medicinal leech: a clinical risk-benefit analysis.

The medicinal leech, Hirudo medicinalis, has been used with increasing frequency for salvage of compromised pedicled flaps and microvascular free-tissue transfers. The data pertaining to 18 cases in which flap salvage with leeches was attempted were reviewed and contrasted with the data from 108 published cases from other centers. A further analysis isolated and examined 19 cases in which infection, secondary to leech utilization, supervened. Results suggested that successful salvage of tissue with leeches occurs in 70 to 80 percent of cases. In contrast, the infection rate of most series is between 7 and 20 percent and, when a clinically significant infection occurs, the flap salvage rate drops to 30 percent or less. In addition to infection, the risks of leech therapy include blood loss, which may require replacement transfusion, loss of leeches into body orifices and spaces, allergic reactions, and adverse psychological responses. Clinical indications and contraindications, and strategies to reduce the risks of leech usage, are discussed. Conclusions are that leeches are best used early, and in accordance with an accepted institutional protocol. For each case, the potential risks and benefits must be rigorously assessed and honestly applied.

Adult↗