The impact of vacuum-assisted excision in the management of indeterminate B3 lesions in the NHS Breast Screening Programme in England.

AIM: To assess the impact of vacuum-assisted excision (VAE) on the management of B3 lesions in the England NHS Breast Screening Programme following an update of national guidance. A secondary aim was to investigate the histological features of malignancies resulting from upgrade of B3 lesions by either VAE or surgery. MATERIALS AND METHODS: The study population was all women recalled for assessment after breast screening who had a wide-bore needle biopsy with a B3 result over the period 01/04/2018 to 31/03/2019. Data were extracted from the National Breast Screening Service (NBSS) computer system at unit level. Women with a B3 result were split into those with and without atypia. The upgrade rates and histological features of malignancies in the different groups were analysed. RESULTS: In total, 2,234,514 women attended for screening between 1/4/218 and 31/3/2019, 84,559 women were referred to assessment, and of those 40,037 women had a core biopsy resulting in 3,355 were B3 lesions (8.38%). Within these, 556 cancers were diagnosed, giving an upgrade rate of 16.57% (556/3,355). The upgrade for B3 lesions with atypia was significantly higher than for B3 lesions without atypia (29.1% versus 13.3%, p<0.001). CONCLUSION: The introduction of the new B3 guidelines has resulted in 73.8% of B3 lesions with atypia and 65.1% of B3 lesions with no atypia having VAE rather than surgery. The data highlights the importance of managing these indeterminate lesions appropriately with an overall upgrade rate of 16.57%.

Contrast-enhanced spectral mammography improves diagnostic accuracy in the symptomatic setting.

AIM: To assess the diagnostic accuracy of contrast-enhanced spectral mammography (CESM), and gauge its "added value" in the symptomatic setting. MATERIALS AND METHODS: A retrospective multi-reader review of 100 consecutive CESM examinations was performed. Anonymised low-energy (LE) images were reviewed and given a score for malignancy. At least 3 weeks later, the entire examination (LE and recombined images) was reviewed. Histopathology data were obtained for all cases. Differences in performance were assessed using receiver operator characteristic (ROC) analysis. Sensitivity, specificity, and lesion size (versus MRI or histopathology) differences were calculated. RESULTS: Seventy-three percent of cases were malignant at final histology, 27% were benign following standard triple assessment. ROC analysis showed improved overall performance of CESM over LE alone, with area under the curve of 0.93 versus 0.83 (p<0.025). CESM showed increased sensitivity (95% versus 84%, p<0.025) and specificity (81% versus 63%, p<0.025) compared to LE alone, with all five readers showing improved accuracy. Tumour size estimation at CESM was significantly more accurate than LE alone, the latter tending to undersize lesions. In 75% of cases, CESM was deemed a useful or significant aid to diagnosis. CONCLUSION: CESM provides immediately available, clinically useful information in the symptomatic clinic in patients with suspicious palpable abnormalities. Radiologist sensitivity, specificity, and size accuracy for breast cancer detection and staging are all improved using CESM as the primary mammographic investigation.

Checkpoint kinase1 (CHK1) is an important biomarker in breast cancer having a role in chemotherapy response.

BACKGROUND: Checkpoint kinase1 (CHK1), which is a key component of DNA-damage-activated checkpoint signalling response, may have a role in breast cancer (BC) pathogenesis and influence response to chemotherapy. This study investigated the clinicopathological significance of phosphorylated CHK1 (pCHK1) protein in BC. METHOD: pCHK1 protein expression was assessed using immunohistochemistry in a large, well-characterized annotated series of early-stage primary operable invasive BC prepared as tissue microarray (n=1200). RESULT: pCHK1 showed nuclear and/or cytoplasmic expression. Tumours with nuclear expression showed positive associations with favourable prognostic features such as lower grade, lower mitotic activity, expression of hormone receptor and lack of expression of KI67 and PI3K (P<0.001). On the other hand, cytoplasmic expression was associated with features of poor prognosis such as higher grade, triple-negative phenotype and expression of KI67, p53, AKT and PI3K. pCHK1 expression showed an association with DNA damage response (ATM, RAD51, BRCA1, KU70/KU80, DNA-PKCα and BARD1) and sumoylation (UBC9 and PIASγ) biomarkers. Subcellular localisation of pCHK1 was associated with the expression of the nuclear transport protein KPNA2. Positive nuclear expression predicted better survival outcome in patients who did not receive chemotherapy in the whole series and in ER-positive tumours. In ER-negative and triple-negative subgroups, nuclear pCHK1 predicted shorter survival in patients who received cyclophosphamide, methotrexate and 5-florouracil chemotherapy. CONCLUSIONS: Our data suggest that pCHK1 may have prognostic and predictive significance in BC. Subcellular localisation of pCHK1 protein is related to its function.

Qualitative mammographic findings and outcomes of surveillance mammography after partial breast reconstruction with an autologous flap.

BACKGROUND: This study describes the qualitative mammographic features after partial breast reconstruction with an autologous flap, and evaluates the diagnostic accuracy and recall rates of surveillance mammography after volume replacement in breast conserving surgery. METHODS: Patients who had autologous partial breast reconstruction (N = 102) after breast-conserving surgery using either the myocutaneous latissimus dorsi mini-flap (N = 39) or fasciocutaneous chest wall perforator flap (N = 63) were reviewed. Mammograms done at one-year post surgery were analysed for characteristic qualitative features. All surveillance mammograms, diagnostic imaging and medical records were retrospectively reviewed. RESULTS: Mammograms of partially reconstructed breasts had distinctive features that correlated well with the surgical procedures. Median follow-up was 3 years, range 0-11 years. Of 295 surveillance mammograms, six (2%) resulted in a recall for further imaging and 3 (1%) proceeded to needle biopsy. Diagnostic imaging was performed for 13 (13/102, 12.7%) patients with symptoms, and only one (1/102, 1%) required a diagnostic biopsy. CONCLUSIONS: A conserved breast with an autologous flap within has characteristic mammographic features that differ from those after standard breast conserving surgery. Surveillance mammography after partial breast reconstruction is accurate, and recall/biopsy rates are low. Diagnostic breast ultrasound examination is effective evaluation for the symptomatic patient.

Optimal screening mammography reading volumes; evidence from real life in the East Midlands region of the NHS Breast Screening Programme.

AIM: To assess real-life reader performance as a function of both volume of mammograms read and reading experience in a multicentre cohort. MATERIALS AND METHODS: Thirty-seven film readers from the East Midlands Breast Screening Programme had 3 years of consecutive screen reading results available for comparison. Markers of screening performance as the first film reader [cancer detection rates, small cancer detection rates, recall rates, positive predictive value of recall (PPV), and missed cancers] were compared with both volume of films read and years of film reading experience. For reading volume, readers were categorized according to film reading volume over the 3 year period: < 15,000 (i.e., on average less than the recommended 5000/year); 15-< 20,000; 20-< 25,000; and ≥ 25,000. For years of experience, readers were categorized into the following groups: < 5 years, 5-< 10 years, 10-< 15 years, and 15-< 20 years. RESULTS: There was no evidence to suggest a relationship between years of film reading experience and film-reading performance. For reading volume, there was a significant difference in the distribution of cancer-detection rate between the four groups (p=0.01); however, there was no difference in small cancer-detection rates, missed cancers or PPV. The median cancer detection rate in the high-volume group (≥ 25,000 mammograms/3 years) was significantly lower than the other groups combined (6.9 per 1000 women screened versus 7.9 per 1000 women screened). The lowest median recall rate was also in the high-volume group, whilst those readers not meeting the NHSBSP minimum requirement had the highest median recall rate; however, there was borderline evidence to suggest a difference in the distribution of recall rates between the four groups. CONCLUSION: The data from the East Midlands do not provide any evidence for reducing the threshold volume of 5000 cases /year. However, there appears to be an upper limit above which reader performance deteriorates in terms of lower cancer-detection rates. With the imminent expansion of the programme, this has implications for service quality. These higher-volume readers should be encouraged to recall more borderline cases to assessment. Analysis of national data is recommended to further evaluate these findings.

A survey of current UK practice regarding the biopsy of clinically and radiologically benign breast masses in young women.

AIM: To determine current practice in the UK of needle sampling of clinically and radiologically benign breast masses in young women. METHOD: A questionnaire regarding needle sampling practice in young women with clinically and radiologically benign breast masses was sent to 481 members of the Royal College of Radiologists Breast Group. This included questions on whether a written protocol is in place to allow avoidance of biopsy, and if so, the clinical and radiological criteria used. RESULTS: Responses were available for analysis from 80 units. Forty-two (53%) units had no written policy in place, whilst 38 (47%) adhered to a written policy. Of those with a policy, an age criterion for safe avoidance of biopsy was present in 36 out of the 38 units (95%). The age limit used ranged from <25 years to <35 years. Twenty-seven (71%) written policies included clinical criteria but only four (10%) policies included a size criterion. Radiological criteria were present in all policies and the majority (74%) adhered to full Stavros criteria. Seven units (18%) used a revised form of the Stavros criteria and three units used their own criteria. CONCLUSION: There is little concordance between symptomatic breast clinics regarding the criteria for avoidance of breast biopsy in this young patient group. Given the very low incidence of breast carcinoma in women less than 25 years old it is considered safe and feasible to adopt a standardized protocol across the UK and avoid the often-unnecessary benign biopsies in these patients.

Does the insertion of a gel-based marker at stereotactic breast biopsy allow subsequent wire localizations to be carried out under ultrasound guidance?

AIM: To investigate whether the insertion of a gel-based marker at the time of stereotactic breast biopsy allows subsequent preoperative localization to be performed under ultrasound guidance. MATERIALS AND METHODS: One hundred consecutive women who underwent either a 10 G stereotactic vacuum-assisted breast biopsy or 14 G stereotactic core biopsy with marker placement, followed by wire localization and surgical excision were identified. All had mammographic abnormalities not initially visible with ultrasound. The method of preoperative localization was recorded and its success judged with reference to the wire position on the post-procedure films relative to the mammographic abnormality and the marker. Histopathology data were reviewed to ensure the lesion had been adequately excised. RESULTS: Eighty-three women (83%) had a successful ultrasound-guided wire localization. Successful ultrasound-guided localization was more likely after stereotactic vacuum biopsy (86%) compared to stereotactic core biopsy (68%), although this did not quite reach statistical significance (p=0.06). CONCLUSION: The routine placement of a gel-based marker after stereotactic breast biopsy facilitates preoperative ultrasound-guided localization.

Does computer-aided detection have a role in the arbitration of discordant double-reading opinions in a breast-screening programme?

AIMS: To investigate whether a computer-aided detection (CAD) system could act as an arbitrator of discordant double-reading opinions, replacing the need for an independent third film reader. METHODS: The mammograms of the 240 women that underwent arbitration by an independent third reader were identified from the 16,629 women attending our screening centre between July 2003 and April 2004. Mammograms of the arbitration cases were digitized and analysed by a CAD system. To assess the ability of CAD to act as the arbitrator, the site of the CAD prompts was retrospectively compared to the site of any abnormality noted by the original film readers. If a CAD prompt was placed on a region marked by one of the film readers then the decision of CAD as the arbitrator was that the women should be recalled for further assessment. If no mark was placed then the region was considered low risk and the decision was not to recall. The decision of CAD as the arbitrator was retrospectively compared with the original recall decision of the independent third reader. RESULTS: There were 21 cancer cases in the group of women undergoing arbitration, diagnosed both at the original screening episode and subsequently. The independent third reader recalled 15/18 (83%) of the cancers that corresponded with the arbitrated lesion. CAD as the arbitrator would have recalled 16/18 (89%) of the cancers that corresponded to the arbitrated lesion. CAD acting as the arbitrator would have resulted in a significant increase in normal women being recalled to assessment in the arbitration group (P<0.001). The extra 50 recalls would have potentially increased the overall recall rate to assessment from 3.1 to 3.4%; a relative increase of 10%. CONCLUSIONS: The main effect of CAD acting as an arbitrator of discordant double-reading opinions is to increase the recall rate, significantly above what is found when arbitration is performed by an independent third reader. Using CAD as an arbitrator may be an option to deal with discordant double-reading opinions when no other method of consensus or arbitration is available.

CT staging of loco-regional breast cancer recurrence. A worthwhile practice?

AIMS: To assess the usefulness of computed tomography of the chest, abdomen, and pelvis (CTCAP) in the detection of metastatic disease in patients presenting with loco-regional recurrence of breast cancer, and to identify subgroups particularly likely to have metastases. MATERIALS AND METHODS: Over a 32-month period, 63 patients with 65 recurrences underwent CTCAP, and were identified from the hospital's computerized radiology system. RESULTS: Twenty-one patients (32%) had metastases, including bony (n=5, 8%), liver (n=7, 11%), and thoracic disease (n=11, 17%). Patients with recurrence in a conserved breast had lower rates of metastasis on CT than those with other sites of recurrence [three of 21 (14%) versus 18 of 44 (41%), p=0.03]. Patients younger than 50 years at primary diagnosis or younger than 60 years at recurrence had statistically significantly higher rates of metastasis than older patients [10 of 16 (63%) versus 11 of 48 (23%), p=0.003, and 13 of 23 (57%) versus eight of 42 (19%), p=0.002, respectively]. CONCLUSION: CT staging of patients presenting with loco-regional recurrence of breast cancer is a worthwhile practice. Younger patients and those with recurrence other than in the conserved breast are particularly likely to have metastatic disease.

Can computerised tomography replace bone scintigraphy in detecting bone metastases from breast cancer? A prospective study.

BACKGROUND: The aim of this study was to determine whether bone scans (BS) can be avoided if pelvis was included in CT thorax and abdomen to detect bony metastases from breast cancer. MATERIALS AND METHODS: Results of 77 pairs of CT (thorax, abdomen, and pelvis) and BS in newly diagnosed patients with metastatic breast cancer (MBC) were compared prospectively for 12 months. Both scans were blindly assessed by experienced radiologists and discussed at multidisciplinary team meetings regarding the diagnosis of bone metastases. RESULTS: CT detected metastatic bone lesions in 43 (98%) of 44 patients with bone metastases. The remaining patient had a solitary, asymptomatic bony metastasis in shaft of femur. BS was positive in all patients with bone metastases. There were 11 cases of false positive findings on BS. CONCLUSION: Our findings suggest routine BS of patients presenting with MBC is not required if CT (thorax, abdomen, and pelvis) is performed.

Rapid distribution of tryptophol (3-indole ethanol) to the brain and other tissues.

Tryptophol (3-indole ethanol) is a compound which induces sleep, and is formed: (a) in the liver after disulfiram treatment, and (b) by the parasite in trypanosomal sleeping sickness. We prepared, purified, and characterized radiolabeled tryptophol for the purpose of defining its tissue distribution in animals. Tryptophol was found to be highly lipophilic, with an octanol:water partition coefficient of 29.8. Brain extraction, determined after intracarotid injection, was high (brain uptake index = 117 +/- 3.5%), and nonsaturable, suggesting the absence of a carrier system. After intravenous administration, tryptophol distribution to tissues correlated with relative blood flow. More than 85% of the radioactivity remaining in brain 2-5 min after intravenous injection co-migrated with tryptophol standards when analyzed by thin-layer chromatography. Other evidence suggested that tryptophol binds to serum and in vivo may be stripped from serum albumin and taken up by brain in a single capillary transit. Our study suggests that in states such as trypanosomal sleeping sickness or disulfiram treatment, remotely formed tryptophol gains ready access to brain (it is 100% cleared in a single capillary passage), and could thus cause somnolence.

Melphalan penetration of the blood-brain barrier via the neutral amino acid transporter in tumor-bearing brain.

Melphalan, a nitrogen mustard derivative of the neutral amino acid L-phenylalanine, was transported across the rat blood-brain barrier by the large (L-system) neutral amino acid transporter in tumor-bearing brain, but no evidence for blood-brain barrier transport by the alanine-serine-cysteine system carrier was obtained in the present study. The ability of melphalan to inhibit phenylalanine uptake was compared in rats implanted with two experimental CNS tumors: the C-6 glioma (a model of primary brain tumors) and Walker carcinoma (a model of metastatic brain tumors). The melphalan concentration which caused 50% inhibition of blood-brain barrier (BBB) phenylalanine uptake (Ki) was 0.49 +/- 0.18 mM in the Walker tumor, compared with 0.46 +/- 0.19 mM in the contralateral control brain. In the ipsilateral hemisphere (Ki = 0.59 +/- 0.25 mM) and contralateral hemisphere (Ki = 0.45 +/- 0.19 mM), drug entry was also via the neutral amino acid transporter. In C-6 gliomas (Ki = 0.77 +/- 0.20 mM) and contralateral control brain (Ki = 0.84 +/- 0.29 mM), melphalan also inhibited BBB phenylalanine transport. A major finding was that, at melphalan concentrations greater than 1.0 mM, BBB permeability of radiolabeled indium (chelated to EDTA) increased in proportion to melphalan concentration. In the contralateral hemisphere of rats implanted with C-6 gliomas, brain extractions of indium-EDTA measured 3 to 4% in the absence of drug, 5 to 6% at 2.5 mM melphalan, and 9 to 10% at 5 mM melphalan. A similar phenomenon was observed in the nontumoral brain regions of rats implanted with Walker carcinoma cells. In normal (nonimplanted) rats, melphalan's inhibition (Ki = 0.29 mM) of phenylalanine and tryptophan (Ki = 0.20 mM) uptake was confirmed, and brain extraction of sucrose (a nonspecific marker which does not penetrate the intact BBB) was observed to increase in proportion to melphalan concentration. We conclude that melphalan not only enters the brain via the neutral amino acid transporter, but at higher concentrations (greater than 1 mM) may open the blood-brain barrier in a nonspecific manner.

Independent blood-brain barrier transport systems for nucleic acid precursors.

The blood-brain barrier permeability to certain 14-C-labelled purine and pyrimidine compounds was studied by simultaneous injection in conjunction with two reference isotopes into the rat common carotid artery and decapitation 15s later. The amount of 14-C-labelled base or nucleoside remaining in brain was expressed in relation to 3-H2O (a highly diffusible internal standard) and 113m-In-labelled EDTA (an essentially non-diffusible internal standard). Of the 17 compounds tested, measurable, saturable uptakes were established for adenine, adenosine, guanosine, inosine and uridine. Two independent transport systems in the rat blood-brain barrier were defined. One transported adenine (Km equals 0.027 mM) and could be inhibited with hypoxanthine. Adenosine (Km equals 0.018 mM), guanosine, inosine and uridine all cross-inhibit, defining a second independent nucleoside carrier system. Adenosine inhibited [14-D]uridine uptake more effectively than did uridine, suggesting a weaker affinity of uridine for this nucleoside carrier.

A comparison of total body and local spinal cord irradiation in experimental allergic encephalomyelitis.

Spinal cord distribution spaces of 3H-mannitol (M.W. 182) were measured to compare the effects of both total-body gamma irradiation and local (spinal cord) x-irradiation on the development of experimental allergic encephalomyelitis (EAE). Single doses of 700R, 500R, 300R, 200R and 100R total body gamma irradiation, administered 6-8 days after EAE induction, appeared to delay the onset, reduce the severity, and hasten recovery of the disease. The magnitude of this effect was approximately prportional to the administered radiation dose. A single 1000R x-irradiation dose, delivered to a 1 x 2cm area in the thoracolumbar region of the spinal cord 7 or 10 days after induction appeared to increase the severity of EAE. Mannitol distribution spaces were mildly elevated in the thoracic regions and significantly elevated in the lumbar spinal cords of cord-irradiated animals. A marked exacerbation of the disease was observed in cord-irradiated rats treated with 5000R, administered over a period of 5 days. Rats progressively lost weight during the normal course of EAE, and a simultaneous increase in wet weights of the spinal cords was seen. Slight changes in these two parameters were observed in cord-irradiated, but not in total-body irradiated animals.

Glut1 glucose transporter in the primate choroid plexus endothelium.

The objective of the present study was to define the cellular location of the Glut1 glucose transporter in the primate choroid plexus. Immunogold electron microscopy indicated that Glut1 epitopes were associated primarily with choroid plexus endothelial cells. Digitized analyses of electron microscopic images provided quantitative estimates of the relative number of Glut1 glucose transporter epitopes on luminal and abluminal endothelial cell membranes within the choroid plexuses. We recorded a high density of Glut1 in the microvascular endothelium of primate choroid plexus, which was consistent in vervet monkeys (5-10 Glut1 gold particles per micrometer of endothelial cell plasma membrane), as well as in baboons (5-20 Glut1 gold particles per micrometer of capillary plasma membrane). In the baboon choroid plexus, we observed that perivascular cells (presumed to be pericytes) were also Glut1-positive, but with substantially reduced activity compared with endothelial cells. Occasional Glut1-immunogold particles were also seen in the basolateral membranes of the choroid plexus cuboidal cells. Light microscopic immunocytochemistry confirmed the abundance of Glut1 immunoreactivity in choroid plexus endothelial cells of vervet monkeys and baboons. A similar pattern was observed in surgically resected human choroid plexus, suggesting differences between primates, including humans and laboratory animals. The only difference was that erythrocytes within the human choroid plexus exhibited a florid Glut1-positive response, but were weakly immunoreactive in nonhuman primates. The observation of high glucose transporter densities in choroid plexus endothelial cells is consistent with the suggestion that choroidal epithelia and capillaries provide a metabolic work capability for maintaining ionic gradients and secretory functions across the blood-CSF barriers.

A single glucose transporter configuration in normal primate brain endothelium: comparison with resected human brain.

Cellular distribution of the Glut1 glucose transporter in normal primate brains was analyzed by immunogold electron microscopy. Two configurations of endothelial Glut1 glucose transporter (high and low density capillaries) have been found in resections of traumatically injured and epileptogenic human brain; the objective of the present study was to ascertain whether these same 2 capillary populations, expressing high and low glucose transporter densities, were the common configuration in normal brain. The relative numbers of Glut1 glucose transporter-associated gold particles on luminal and abluminal endothelial cell membranes were determined within the cerebral cortex of several normal, nonhuman primates. Low Glut1 densities were seen in brain endothelia of both the rhesus and squirrel monkey cortex, with slightly greater quantities of Glut1 in vervet monkey cortices. The Glut1 transporter was most highly expressed in the baboon cortex, approaching the concentrations seen in human brains. In the rhesus, squirrel, and vervet monkeys, Glut1 concentrations were greater on the abluminal than luminal capillary membranes. In contrast, mean luminal membrane Glut1 concentrations were greater in baboons, resembling the distribution seen in the human brain. Brain regional differences in transporter concentration were seen in comparing membrane densities in the baboon cortex (approximately 15 Glut1-gold particles per micrometer), hippocampus (approximately 12 Glut1 gold particles per micrometer), cerebellum (approximately 6 Glut1-gold particles per micrometer), and retinal microvasculature (approximately 20 Glut1-gold particles per micrometer). We conclude that a single, uniform Glut1 distribution characterizes brain capillaries of normal nonhuman primates, and hypothesize that the presence of high and low density glucose transporter endothelial cells (seen in human traumatic injury and seizure resections) represents a pathologic response to brain insult.

High expression of the Glut1 glucose transporter in human brain hemangioblastoma endothelium.

The principal glucose transporter at the blood-brain barrier is Glut1, and GLUT1 expression is downregulated in high grade gliomas. In the present study, glucose transporter expression was studied in surgically resected hemangioblastoma tissue. Light microscopic immunochemistry indicated the high expression of the Glut1 glucose transporter isoform throughout the central vascular endothelium of this tissue. Glial fibrillary acidic protein (GFAP) was observed only at the tumor border, with no GFAP immunoreactivity in stromal cells, pericytes or endothelia in the central tumor regions. It is generally believed that more Glut1 is found in erythrocytes than any other cell, but quantitative electron microscopic immunogold analyses of Glut1-immunoreactive sites per micron of capillary membrane showed the Glut1 density in tumor endothelial membranes glucose transporter was 2-3-fold higher than in human red cells. In the same tissue samples, qualitative immunogold electron microscopy of human serum albumin indicated that this protein (MW 65,000) moved freely from the vascular space into pericapillary regions, confirming the leaky barrier characteristics of the hemangioblastoma. These studies show that Glut1 expression may be high in endothelia that are highly permeable and devoid of astroglial contacts. Thus, human cerebral hemangioblastomas may provide a novel system for studying the induction of Glut1 in the blood-brain barrier.

Blood-brain barrier restriction of peptides and the low uptake of enkephalins.

Blood-brain barrier penetration of leucine-enkephalin, methionine-enkephalin, and other peptide-like compounds was measured after intracarotid injection of three isotopes and was found to be non-saturable over the nanomolar range of concentrations tested. No significant differences in brain regional extraction of leucine enkephalin (or morphine or heroin) were observed. In contrast to previous reports, the brain extraction of enkephalins was minimally low (E = 2-3%) and about the same order of magnitude as other putative neurotransmitters. Brain extractions of other peptide-like compounds were similarly small: TRH, E = 1%; glutathione, E = 0.5%; beta-alanyl histidine, E = 1%; and thioacetyl coenzyme A, E = 2%. Extraction of the non-diffusible reference dextran was determined to be 1%, suggesting that the blood brain barrier tends to restrict peptide penetration.