ECMUS Literature Reviews

Kieler H, Haglund B, Cnattingius S, Palmgren J, Axelsson O. Does prenatal sonography affect intellectual performance? Epidemiology 2005; 16: 304-310

This is a registry study of a possible association between exposure to ultrasound during pregnancy and intellectual performance among 18 years old males.

The information about intellectual abilities was retrieved from the Swedish National Service Register which includes the results of a battery of tests the men are submitted to when enrolling for military service at the age of 18 years. The test results are given as standard scores (maximum 9, mean 5, SD 2).

Data about the place of birth of a cohort of more than 200,000 individuals were available from the Swedish Medical Birth Register. After exclusions, two groups were compared - a group of 6026 men, supposedly exposed to ultrasound as fetuses, and a control group of 161,033 men. The exposure to ultrasound was assumed to have happened as the men of the index group were born in the Southern Swedish area served by the Malmö Department of Obstetrics and Gynecology that was the only unit in Sweden that offered an ultrasound examination as a routine for detection of multiple pregnancies and dating to all pregnant women during the years 1973-1978. Two additional comparisons were performed: the results of the index cohort were compared to the results of the period before introduction of routine ultrasound in Malmö and analyses were also performed on 15,540 pairs of brothers. The older brothers were born before, and the younger brothers after, the introduction of ultrasound examinations; in 456 of the pairs, the younger brothers were born in Malmö.

The men born in Malmö were found to have lower intellectual performance scores (mean difference -0.16; 95 % confidence interval [CI] -0.21 to -0.11) and an increased risk of subnormal performance (odds ratio [OR] 1.28; CI 1.18.1.38) as compared with the control group. However, when analysing the period before the introduction of ultrasound examinations, the men born in Malmö also had lower intellectual performance and the difference was of similar magnitude. In addition, there were no differences within brother pairs, when comparing the younger brothers, assumed to have been exposed to ultrasound, and the unexposed older brothers. Thus, the authors concluded that there was no association between ultrasound examination during pregnancy and intellectual performance in young adults.

The present study is large, and the information was collected prospectively in the registries. Nevertheless the study has limitations; possible confounding factors are largely unknown, the intellectual test used at enrolment may be questioned, and the exposure information is assumed, based only on information about the place of birth. This is adequately discussed in the paper. The results do not suggest a negative influence of ultrasound examination in pregnancy on the intellectual development in males.

Stålberg K, Haglund B, Axelsson O, Cnattingius S, Hultman CM, Kieler H. Prenatal ultrasound scanning and the risk of Schizophrenia and other psychoses. Epidemiology 2007; 18: 577-82

Prenatal ultrasound exposure has been associated with increased prevalence of left-hand or mixed-hand preference and has been suggested to affect the normal lateralization of the fetal brain. Atypical lateralization is more common in patients with schizophrenia. Thus, a study on the possible association between prenatal ultrasound and schizophrenia is welcomed.

This cohort study assesses ultrasound exposure from aggregate data. The University Hospital in Malmö was the first hospital in Sweden to use ultrasound scans as part of standard antenatal care. Children born in Malmö in 1973-78 were considered exposed to ultrasound. They were compared with children born in 1973-78 at other Swedish hospitals that had not yet introduced ultrasound. Those children were considered unexposed to ultrasound. The Swedish Medical Birth Register (information on place of birth) was linked to the Hospital Discharge Register and the Cause of Death Register.

In all, 370,945 individuals were included in the study, of whom 13,212 were assumed to have been exposed to ultrasound.

ECMUS Literature ReviewsThe exposed group demonstrated a tendency toward a higher risk of schizophrenia (crude incidence rate among men = 1.58,95 % CI 0.99-2.51, and among women = 1.26,95 % CI 0.62-2.55). However, men and women born in several other tertiary level hospitals without ultrasound scanning facilities also had higher risks of schizophrenia compared with those born in other hospitals. For other psychoses there were no differences between groups. The authors concluded that there were no clear associations between prenatal ultrasound exposure and schizophrenia or other psychoses. Other factors related to place of birth might have influenced the results.

Comment

The authors found a borderline statistically significant association in men. Being a cohort study, there might be bias or confounding factors involved in explaining the association. There is no doubt that the simplistic method of assigning ultrasound exposure (on the basis of place of birth) has misclassified some of the individuals in the study. The authors validly argue that this misclassification bias will lead to an underestimation of the association and reduce the possibility to find an existing relation between exposure and outcome.

This is an important study with a well-founded conclusion, that there is probably no association between prenatal ultrasound and schizophrenia or other psychoses later in life. However, the present study addressed US devices in clinical use in 1973-1978. It is a well-known fact that intensity output levels from modern devices may be 10-15 times higher. Thus, the study may be of limited relevance for current obstetric ultrasound scanning.

Vykhodtseva N, McDannold N, Hynynen K. Progress and problems in the application of focused ultrasound for blood-brain barrier disruption. Ultrasonics. 2008; 48: 279-96

Do you know what the major feature of the blood brain barrier is? The endothelial cells of brain capillaries are uniquely bound to their neighbours by tight junctions which restrict the passage of drugs into the brain to small molecules and some lipophilic drugs such as narcotics. No other drugs can enter the brain and reach a site of action. Two methods have succeeded in disrupting the blood brain barrier over the last decades, intra-arterial infusion of hyperosmolar mannitol and focused ultrasound, yet these have never progressed beyond an initial experimental stage. Some time ago, researchers in Boston found a new way of disrupting the blood brain barrier locally by adding contrast agent to ultrasound. They have recently summarised the current state of their research in a review article [1]. The major findings were, in brief:

1. Ultrasound disrupted the blood brain barrier transiently. The barrier was open for one or 2 hours and was closed again 4 hours after sound application.

2. Morphological examination of the brain showed little damage with only few focal extravasations of red blood cells and no damage to neurons. This contrasted to previous experiments where haemorrhage with focal neuronal death had been found.

3. A standard animal experimental system was established to allow investigation of blood brain barrier opening. To facilitate sound entry through the skull, a piece of skull bone was removed and rabbit and rat brains were exposed through a cranial window. Magnetic resonance imaging with an otherwise impermeable gadolinium containing contrast agent delineated regions where the blood brain barrier was open.

4. Experiments on the impact of ultrasound exposure parameters on blood brain barrier opening revealed that

   - a Lower frequencies opened the blood brain barrier more efficiently. When 260 and 690 kHz were compared with 1.62 and 2.07 MHz, larger areas of local blood brain barrier opening were found with the 2 low frequencies. In addition the number of extravasated red blood cells was lower at the low frequencies.

   - b The pressure thresholds for producing blood brain barrier opening at these frequencies rose accordingly from 0.25 MPa at 260 kHz to 0.69 MPa at 2.1 MHz.

   - c Determination of the mechanical index (MI) at these pressures and frequencies revealed a threshold MI of 0.46 where blood brain barrier disruption began [1].

   - d Rabbit brains were originally exposed to 20 ultrasound bursts of 10 or 100 ms duration at 1 Hz . Shortening the burst length to 1 and 0.1ms produced an inferior result [2]. Doubling the speed of burst application from 1 Hz to 2 Hz made no difference.

5. 5. A closer look at the ultrastructure of tight junctions in the brain revealed that their molecular structure was transiently disassembled [3]. Antibodies against the tight junction proteins occludin, claudin-5 and ZO-1 produced less staining during disassembly one and 2 hours after sonication. In addition a marker protein passed through endothelial cells and also along open endothelial clefts. Four hours after ultrasound the tight junctions were reassembled.

6. 6. As to the size of molecule able to pass the blood brain barrier, small molecules such as trypan blue and contrast agents passed, as did large molecules such as 40 kDa peroxidise, 180 kDa antibodies and nanoparticles.

7. 7. To demonstrate the potential to deliver a drug locally into the brain, the monoclonal antibody Herceptin, a humanized anti-human epidermal growth factor receptor 2 monoclonal antibody, was transferred at 260 kHz ultrasound into rabbit brains [4]. Immunostaining verified its localisation in endothelial cells.

In summary, a new type of access to the blood brain barrier is described, causing minimal histological damage and enabling the transfer of drugs across it. This method may represent a powerful technique for the delivery of macromolecular agents such as antibodies to treat patients with diseases of the central nervous system.

References

1. McDannold N, Vykhodtseva N, Hynynen K. Blood-brain barrier disruption induced by focused ultrasound and circulating preformed microbubbles appears to be characterized by the mechanical index. Ultrasound Med Biol. 2008; 34: 834-840.

2. Dannold N, Vykhodtseva N, Hynynen K. Effects of acoustic parameters and ultrasound contrast agent dose on focused-ultrasound induced blood-brain barrier disruption. Ultrasound Med Biol. 2008; 34(6): 930-937.

3. Sheikov N, McDannold N, Sharma S, Hynynen K. Effect of focused ultrasound applied with an ultrasound contrast agent on the tight junctional integrity of the brain microvascular endothelium. Ultrasound Med Biol. 2008; 34: 1093-1104.

4. Kinoshita M, McDannold N, Jolesz FA, Hynynen K. Noninvasive localized delivery of Herceptin to the mouse brain by MRI-guided focused ultrasound-induced blood-brain barrier disruption. Proc Natl Acad Sci U S A. 2006 Aug 1; 103(31): 11719-1723.

Williams AR, Wiggins RC, Wharram BL, Goyal M, Dou C, Johnson KJ, Miller DL. Nephron injury induced by diagnostic ultrasound imaging at high mechanical index with gas body contrast agent. Ultrasound Med Biol. 2007; 33: 1336-1344

Ultrasound contrast agents are micrometre sized gas bubbles with a phospholipid or protein shell which can pass through capillaries without getting stuck. Their use for diagnostic imaging has increased explosively in the last years and has started to replace more expensive CT and MRI investigations. As it passes through a capillary, ultrasound excites a gas bubble to resonate; its diameter enlarges and the shell breaks up. When this happens in a capillary, its wall may be torn and red blood cells spill over into the interstitial space generating petechial haemorrhage. In a previous literature review, up to 40 % of the surface of rat kidneys was covered with petechial haemorrhages when ultrasound was applied at a diagnostic level [1]. Important questions were investigated such as the dependence of petechiae on MI, frequency, frame rate, insonation time, and microbubble dose.

Renal haemorrhage induced by ultrasonic contrast agent has recently been re-examined in a study in which 30 rat kidneys were exposed for one minute to 1.5 MHz ultrasound during contrast agent infusion. The organ was examined a short time after treatment and additionally one, 2 or 3 days later. Shortly after treatment, the renal surface was covered with petechiae as expected, yet kidneys examined one or more days later had fewer petechiae - so where was the blood? It had simply drained out of the glomerulus and drained via the tubular system. Urine samples of treated rats confirmed microhaematuria. Histology showed glomerular fibrin deposits, focal cell necrosis and red blood cell and hyaline casts in tubuli, a picture well known from treatment with extracorporeal shock waves for renal stone fragmentation.

A good outcome, sonographers and newsletter readers might suppose, glomerular damage will be repaired and the integrity of the nephron with its blood supply will be restored. Yet this conclusion might be grossly misleading. The experiment's shortcoming is that it did not examine whether glomerular haemorrhage was repaired. The alternative is that glomerular haemorrhage is an irreversible event, the nephron dies and glomerulosclerosis ensues. This is known from other forms of intervention, along with microhaematuria. Experiments involving longer observation periods, sequential studies with histological documentation and adequate animal numbers are needed to clarify whether haemorrhage into Bowman's space is a repairable accident and if nephron function is restored or the renal filtration unit is lost.

The question whether haemorrhage is repaired has implications for the future use of ultrasound contrast imaging in the kidney. Multiple examinations with ultrasound contrast agent in a patient with pre-existing renal disease might aggravate his disease.

1. EFSUMB Newsletter September 2003 page7: Wible JH, Galen KP, Wojdyla JK, Hughes MS, Klibanov AL, Brandenburger GH. Microbubbles induce renal hemorrhage when exposed to diagnostic ultrasound in anaesthetised rats. Ultrasound Med Biol. 2002 Nov-Dec;28 (11-12): 1535-46.