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Effect of Different Pharmacologic and Chemical Agents on the Integrity of Hydatid Cyst Membranes

Mustafa Karaolanolu¹, Ömer Faruk Akinci², Sedat Bozkurt¹, eyhmus Deniz¹, Gülhiz Karata¹, Ali Cokun² and Salih Zeki Ziylan¹

¹ Radiology Department, Harran University Medical Faculty, anliurfa 63100, Turkey.
² General Surgery Department, Harran University Medical Faculty, anliurfa 63100, Turkey.

Received February 24, 2003; accepted after revision November 25, 2003.

 
Presented at the 2003 National Radiology Congress, Ankara, Turkey.

Address correspondence to M. Karaolanolu, Emniyet Caddesi, Bahadir sokak, Ilkin apartmani, A Blok, No. 8, anliurfa 63200, Türkiye.

Abstract

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OBJECTIVE. We performed an in vitro investigation of the effects of widely used scolicidal and sclerosing agents, as well as some pharmacologic products, on the integrity of the membrane of hydatid cysts.

MATERIALS AND METHODS. Two milliliters each of 22 agents, 2 mL of clear fluid, and one piece of hydatid cyst membrane were put into bottles. The hydatid cyst membranes were evaluated by visual observation and manual palpation. Visual examination of the bottles was performed daily for 7 days, and observations of membrane changes, including translucency, destruction, swelling, and melting, were recorded. Manual evaluation was done on the seventh day by finger examination, and membrane fragility was scored.

RESULTS. The hydatid cyst membrane was completely melted in a few minutes in a 2.5% solution of sodium hypochlorite and in 1 hr by a 0.1% sodium hypochlorite solution. The integrity of the hydatid cyst membrane was preserved in alcohol, acetone, glutaraldehyde, albendazole, acetylsalicylic acid, formaldehyde, lidocaine, hydrochloric acid, ammonia, pancreatin, Betadine, methylene blue, and isotonic saline samples. The membranes in the metronidazole and hypertonic saline solutions were not damaged but showed significantly increased fragility. The membranes in levamisole and piperazine hexahydrate became translucent and showed moderate fragility.

CONCLUSION. None of the agents that are used in clinical practice had important effects on the dissolution of hydatid cyst membranes. However, sodium hypochlorite solutions completely melted the hydatid cyst membranes. Because the use of this agent on living tissue is limited, further study is needed to investigate its clinical use.

Introduction

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Percutaneous drainage has been increasingly used as an alternative to surgery in the treatment of hydatid cysts [1-12]. Surgical procedures ordinarily involve inactivation of the cyst contents, then the removal of all cyst components [1, 10]. Percutaneous drainage has many advantages; however, hydatid cyst membranes, which are composed of a laminar layer and a germinative layer, cannot be removed by this method. Percutaneous drainage may be performed by puncture, aspiration of cyst contents, injection of scolicidal agents, and reaspiration of fluid, as described by Ben Amor et al. [13], or by catheterization, as described by Akhan and Özmen [2]. Regardless of which method is used, 5-10 years of follow-up is advocated because of the potential for recurrence or infection [2]. Even after 3 years, membrane fragments may appear sonographically as a pseudotumor [2-4, 9]. Hydatid cyst membranes may limit the effectiveness of percutaneous catheter drainage. In some patients, the needle may become obstructed during aspiration as a result of membrane fragments that occlude the needle lumen [11]. If continuing drainage is required, obstruction of the catheter by membrane fragments could be a significant problem [4, 7]. Studies have shown that agents such as alcohol and hypertonic saline do not break hydatid cyst membranes into pieces that can be drained easily from a catheter [14]. The inability to completely drain hydatid cyst contents percutaneously resulted in surgical referral in 6.3% of patients in one series [1].

Örmeci et al. [12] used 1% polidocanol to augment the effect of alcohol to destroy hydatid cyst membranes. Those authors observed less recurrence after percutaneous drainage, suggesting that using polidocanol instead of hypertonic saline may explain this favorable result. A pharmacologic agent that can dissolve hydatid cyst membranes may solve these problems. An agent that melts hydatid cyst membranes without harming the host tissue might increase the effectiveness of percutaneous drainage.

Although we consider the hydatid cyst membrane to be the focal point of some problems related to the treatment of hydatid disease, we have not found conclusive studies about the effects of scolicidal agents on the hydatid cyst membranes. Our study investigates the effects of widely used scolicidal and sclerosing agents, as well as some pharmacologic products, on the integrity of hydatid cyst membranes in vitro.

Materials and Methods

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Our study was designed to investigate the effects of different agents on hydatid cyst membranes. Two milliliters of each agent listed in Table 1, in the form of a pure solution, was put into a 20-mL bottle; 2 mL of clear fluid from the cyst of a patient who was surgically treated for hydatid disease was added to each bottle. The hydatid cyst membrane obtained from the same patient was cut into pieces measuring 2 x 1 cm. One piece was put into each bottle, and the bottles were numbered from 1 to 23. Only clear water was added to the control bottle. Because some of these agents may have toxic effects, the study was carried out in vitro. All pharmacologic or chemical agents used in this study are listed in Table 1.

A researcher evaluated the samples in a blinded fashion by visual observation and by manual examination. Observation was performed at 15, 30, and 60 min; then at 2, 3, 4, 6, 9, 12, and 24 hr; and then at 2, 3, 4, 5, 6, and 7 days. Translucency, destruction, swelling, and melting were checked and noted (Tables 2 and 3). Because no appreciable change was seen between days 2 and 7, these days were not recorded in the tables. Destruction and swelling were graded as "no change," "moderate," or "severe" compared with the control bottle. A hairy appearance or minimal fragmentation was categorized as moderate destruction. Swelling was graded as compared with the control bottle; swelling more than twice the size of the control sample was termed "severe," and swelling less than twice, "moderate." Translucency was categorized as present or absent.

Manual examination was performed only on samples that showed preserved membrane integrity on day 7. The fragility of the membranes was evaluated by finger examination. Membranes were checked in terms of durability against finger pressure and scored as not fragile, moderately fragile, or severely fragile compared with the control sample.

All bottles were kept closed at room temperature. Because the contents of the bottles containing Betadine (povidone iodine, Kim-Pa), methylene blue, and pancreatin were colored, making membrane observation impossible, these samples were evaluated by emptying the contents into a larger container. After evaluation, the membranes and the solutions were returned to the bottles. The final evaluation was performed on day 7.

Records were kept by one of the researchers in a blinded fashion, and observed changes were scored as follows: 1, the integrity of the hydatid cyst membrane was preserved with no melting, destruction, swelling, or translucency; 2, a moderate change occurred in membrane swelling or destruction; 3, a severe change occurred in membrane destruction or swelling, or the membrane became translucent; 4, the membrane was fragmented into pieces smaller than 2 mL; and 5, the membrane melted completely.

Results

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All observed changes and findings on palpation were compared with the control sample. In the control sample, no appreciable changes such as destruction, swelling, translucency, or fragility were noted. Observed changes in the samples through day 7 are shown in Table 2, and the final observation and palpation findings are shown in Table 3.

Sodium hypochlorite solutions at all dilutions dissolved hydatid cyst membranes completely so that no visible trace remained. The elapsed time for complete dissolution was several minutes for the most concentrated (2.5%) sodium hypochlorite solution and 1 hr for the most diluted (0.1%) solution.

The findings in the alcohol and acetone samples were similar. No observable changes were detected for 7 days, and manual examination revealed more rigid and fixed structures on day 7 compared with the control sample. The Betadine and methylene blue samples showed similar results. No visible changes were seen, and the membranes were moderately fragile on day 7 in both samples.

In the albendazole, ammonium, acetylsalicylic acid, glutaraldehyde, hydrogen chloride, isotonic saline, lidocaine, and pancreatin samples, the membrane integrity was preserved, but moderate destruction and moderate swelling were observed. These changes started within the first hour in the ammonium, acetylsalicylic acid, glutaraldehyde, and hydrogen chloride samples and after the first day in the albendazole and the lidocaine samples. On day 7, the cyst membranes in the albendazole and pancreatin samples were moderately fragile, but the other membranes remained strong. The cyst membrane in the hydrogen chloride sample was translucent compared with the control sample.

Moderate destruction and severe swelling were noted in the metronidazole and the hypertonic saline samples on day 7. Both membranes were severely fragile at palpation. Although metronidazole and 3% sodium chloride were the agents that caused maximum swelling and fragility, the integrity of the membranes in both solutions was not damaged. Three-percent sodium chloride showed its maximum effect in the first hour and then remained stable. The effect of metronidazole was similar to that of 3% sodium chloride, but metronidazole showed its maximal effect at the fourth hour.

The membranes in the levamisole and piperazine hexahydrate solutions became completely translucent. Contrary to hydrogen chloride, the translucent activity started at the third hour and continued at an accelerated rate. However, the translucent effect of piperazine hexahydrate was more evident than that of levamisole. The membrane in both samples was moderately fragile to palpation.

Discussion

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The three main approaches for the treatment of liver hydatid cysts are medical treatment, surgery, and percutaneous drainage. Although medical treatment is not effective alone, benzimidazole derivatives are commonly used for disseminated systemic disease, in inoperable cases, and for prophylaxis during surgery and percutaneous drainage [6, 15]. Surgery was the only treatment available before the introduction of antihelmintic drugs, but surgery is associated with considerable rates of morbidity, mortality, and recurrence [1, 8]. Surgical procedures range from simple puncture and aspiration of the cyst to liver resections or even transplantation, but the most common technique is total or partial cystectomy [10].

Recently, percutaneous drainage has become an effective alternative to surgery, particularly in patients with multiple cysts, patients for whom surgery is contraindicated, patients who refuse surgery, pregnant patients, and those who relapse after surgery [1]. Percutaneous drainage offers distinct advantages over surgery. It can be performed as an outpatient procedure and does not require general anesthesia. Additionally, percutaneous drainage is relatively inexpensive and requires a shorter hospitalization [2-7]. Many clinical studies have been published on its effectiveness [2-12]. As of 1999, more than 2,500 percutaneous drainage procedures had been performed in several countries by different teams, with low morbidity (4.1%) and mortality (0.08%) rates [10]. However, percutaneous drainage cannot remove or dissolve hydatid cyst membranes and so cannot cure the disease completely in a few days as surgery can.

Hydatid cysts have strong membranes that cannot be easily dissolved or fragmented by common scolicidal agents. A mature hydatid cyst consists of a thick, nonliving laminated outer layer that surrounds a thin inner layer of germinal tissue and a fluid-filled interior. Survival of the cyst may be aided by the presence of the laminated layer, which grows along with the parasite, increasing its thickness with age. The laminated layer has two major functions: to provide mechanical support and to form a barrier protecting the living part of the cyst. Many chemical agents have been used to kill the living part of the cyst; however, the nonliving laminated membrane is highly resistant to such chemicals [16]. The solid part remains undamaged and cannot be removed by aspiration or percutaneous drainage.

The feasibility of percutaneous drainage of hepatic hydatid cysts depends on the morphologic classification of the cyst as described by Gharbi et al. [17]. Those authors divided hydatid cysts into five sonographic types: type I, a pure fluid collection; type II, a fluid collection with a split wall, which is the cyst membrane detached from the pericyst; type III, a fluid collection with daughter cysts; type IV, a solid collection; and type V, calcification of the cyst wall. Classically, type I, type II, and some type III cysts can be treated percutaneously. Some researchers have reported that some type III cysts that contain nondrainable solid material cannot be treated percutaneously [3, 4, 18].

In the near future, percutaneous drainage may be the treatment of choice. We believe that unsuccessful percutaneous drainage of hepatic hydatid cysts is the result of problems associated with the cyst membrane. These problems may be sorted into four main categories. One, percutaneous drainage cannot be applied to all types of cysts; two, the existence of a nondrainable solid content in the cystic cavity reduces the effectiveness of percutaneous drainage; three, prolonged drainage leads to an increased rate of complications; and four, the inability to effect a rapid decrease in cavity size results in prolonged treatment and expense.

First, the solid content of a cyst is the critical factor in determining if it qualifies for percutaneous drainage. Rendering the solid component of a hydatid cyst more easily drainable will allow many cysts to be drained percutaneously. The nondrainable material in types III and IV cysts is formed by pieces of the cyst membrane. Histologic examination of this solid material reveals it to be laminated [4, 7, 9]. Dissolution of these membranes with pharmacologic agents will probably make these cysts more easily drainable.

Second, the existence of solid material in a cyst reduces the effectiveness of percutaneous drainage. In patients in whom continuing drainage is required, obstruction of a catheter by membrane fragments makes the drainage more difficult and increases its duration. Thus, the dissolution of the membrane may help to transform complex hydatid cysts into simple unilocular cysts, allowing the drainage process to be performed faster and more effectively.

Third, prolonged drainage increases the rate of complications. Blockage of the catheter by membranes not only hinders drainage but also increases the possibility of additional complications. Using a larger catheter to solve these problems negatively affects patient comfort, increases tissue trauma, and causes additional complications such as infection, bleeding, pain, and leakage into the peritoneal cavity [7, 19].

Fourth, incomplete drainage of a hydatid cyst results in a residual mass that is detectable by imaging, which raises the anxiety level of both the patient and the referring clinician because of the increased risk of recurrence. Most studies discuss a pseudotumor appearance on sonography long after aspiration of the cyst [2-4, 9]; this appearance reflects the nondrained parts of the cyst membrane [9].

Agents that are currently used do not dissolve hydatid cyst membranes in a manner that allows efficacious percutaneous drainage via a catheter. A pharmacologic agent that can facilitate the dissolution of cyst membranes may solve these problems. The ideal agent should melt the solid components of the cyst without harming the host tissue. Development of an ideal agent would broaden the indications for and increase the effectiveness of percutaneous drainage.

In this study, we considered several features in choosing a pharmacologic agent; we think that an agent that can disintegrate hydatid cyst membranes has many benefits. Preferred agents for such a purpose are hypertonic saline, Betadine, and alcohol. Also important was choosing a pharmacologic agent that could be used as an antiparasitic agent. For this purpose, albendazole, metronidazole, and levamisole are used. Derivatives of benzimidazole are agents of choice for the medical treatment of hydatid cysts. Albendazole was used in an experimental study and found to be effective [20]. Other points considered in choosing a solution were that it had to contain enzymes, it could be used as a chemical solvent, and it should have either an acidic or a basic nature. However, none of the agents we have used showed the expected effects.

In our study, a solution of hypertonic saline (3%) was found to act on hydatid cyst membranes, causing swelling and making them fragile. However, the effect of hypertonic saline was observed in the first few hours, and then the membrane remained stable. On day 7, the membrane integrity was preserved. A solution of hypertonic saline was found to be the most effective compared with other agents (Betadine and alcohol) used in clinical practice. Metronidazole, an antiparasitic drug, showed effects on the hydatid cyst membrane similar to the effects of hypertonic saline. Piperazine hexahydrate and levamisole induced membrane translucency. Although that is an interesting finding, we do not think that making the membrane translucent has any particular importance. One of the most commonly used agents, Betadine, showed a limited effect on the cyst membrane. Alcohol, which is used widely in clinical studies for sclerosis, and acetone did not change the integrity of the membrane but made it more rigid and fixed in form compared with the control sample. Therefore, alcohol appeared not to be suitable for dissolving the membrane. Karayalçin et al. [14] reported that hypertonic (20%) saline and absolute alcohol had no effect on daughter cysts. In their study, macroscopic examination of daughter cysts showed no change in the integrity of the wall after 1 hr of exposure to hypertonic saline and alcohol.

The most remarkable result in this study was the complete dissolution of hydatid cyst membranes by sodium hypochlorite. A 2.5% solution of sodium hypochlorite melted the hydatid cyst membrane completely in the first few minutes. We then studied more diluted forms of this solution, and the most diluted form (0.1%) caused melting of the membrane in 1 hr. Sodium hypochlorite been used in a patient with nephrolithiasis to sanitize pyogangrenous foci during renal surgery [21]. In another study, sodium hypochlorite was used during endodontic treatment of the tooth canal [22]. Also, 0.6% sodium hypochlorite was studied in cataract surgery for cleaning the lens [23]. Girardo et al. [24] showed that sodium hypochlorite has a strong antiseptic effect on gram-negative micro-organisms. Another study showed similar results for Legionella species [25]. Zanini and Graeff-Teixeira [26] showed that sodium hypochlorite is effective on living larvae and can be used for food decontamination. In fact, sodium hypochlorite has been long used for water sanitation. If sodium hypochlorite is sufficiently studied and proven to be safe for host tissue, it may be a perfect agent as an adjunct to percutaneous drainage of hydatid cysts because of its antiparasitic and membrane dissolution properties. Having the potential to improve the success of percutaneous drainage, sodium hypochlorite may be able to expand the indications for percutaneous drainage of hepatic hydatid cysts. We think that further studies are needed to determine the optimal dose and possible toxic effects of this agent.

In conclusion, unlike surgery, percutaneous drainage cannot extract the hydatid cyst membrane, which may reduce both the short- and long-term efficacy of percutaneous drainage. If an agent can be found that is efficacious for dissolving the hydatid cyst membrane, the effectiveness of percutaneous drainage will increase, complications will decrease, and indications for percutaneous drainage will broaden. In our in vitro study, none of the agents used in clinical practice had significant effects on hydatid cyst membranes. However, sodium hypochlorite was effective in dissolving the membranes. Because the use of this agent on living tissue is limited, further study is needed to investigate its clinical usefulness.

Acknowledgments
 
We thank Ahmet Bozkurt and Emily Öztürk for their help in revising the manuscript grammatically, Özcan Erel for preparing and standardzing of the chemical agents used, and Aynur Isin for helping in the experiments.

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