Cisplatin and radiation therapy in HIV-positive women with locally advanced cervical cancer in sub-Saharan Africa: A phase II study of the AIDS malignancy consortium.

PURPOSE: To determine the feasibility, safety, and tolerability of concomitant chemoradiotherapy administered at standard doses in HIV-infected women with locally-advanced cervical cancer (LACC) receiving antiretroviral therapy (ART). PATIENTS AND METHODS: Eligible participants had HIV infection and untreated, histologically-confirmed, invasive carcinoma of the uterine cervix, FIGO stages IB2, IIA (if tumor >4 cm), IIB, IIIA, IIIB, or IVA and met standard eligibility criteria. Subjects were prescribed 41.4-45 Gy external beam radiation therapy followed by high dose rate brachytherapy concomitant with up to six weekly doses of cisplatin 40 mg/m2 and were followed for 12 months. RESULTS: Sixty-four women were screened at two sites in sub-Saharan Africa, of whom 40 eligible participants were enrolled, for a screening ratio of 1.60. Of the 38 eligible participants who initiated study treatment, 31 (82%) completed treatment. By the 12-month follow-up visit, 7 women had died of disease and 29 of 31 (94%) returned for follow-up. One-year progression-free survival was 76.3% (95% CI, 59.4-86.9%), and did not significantly differ according to stage at entry (p = 0.581). Participant-reported adherence to ART was high; by 12 months, 93% of participants had an undetectable viral load. The most common grade 3 or 4 adverse event was decreased lymphocyte count that affected all treated participants. Non-hematologic serious adverse events were similar to those observed in women with LACC without HIV infection. CONCLUSIONS: The majority of HIV-infected women with LACC can complete concomitant chemoradiotherapy with the same cisplatin dose used in HIV-uninfected women with comparable tolerability and high ART adherence while on treatment.

Insulin/IGF and sex hormone axes in human endometrium and associations with endometrial cancer risk factors.

PURPOSE: Experimental and observational data link insulin, insulin-like growth factor (IGF), and estrogens to endometrial tumorigenesis. However, there are limited data regarding insulin/IGF and sex hormone axes protein and gene expression in normal endometrial tissues, and very few studies have examined the impact of endometrial cancer risk factors on endometrial tissue biology. METHODS: We evaluated endometrial tissues from 77 premenopausal and 30 postmenopausal women who underwent hysterectomy for benign indications and had provided epidemiological data. Endometrial tissue mRNA and protein levels were measured using quantitative real-time PCR and immunohistochemistry, respectively. RESULTS: In postmenopausal women, we observed higher levels of phosphorylated IGF-I/insulin receptor (pIGF1R/pIR) in diabetic versus non-diabetic women (p value =0.02), while women who reported regular nonsteroidal anti-inflammatory drug use versus no use had higher levels of insulin and progesterone receptors (both p values ≤0.03). We also noted differences in pIGF1R/pIR staining with OC use (postmenopausal women only), and the proportion of estrogen receptor-positive tissues varied by the number of live births and PTEN status (premenopausal only) (p values ≤0.04). Compared to premenopausal proliferative phase women, postmenopausal women exhibited lower mRNA levels of IGF1, but higher IGFBP1 and IGFBP3 expression (all p values ≤0.004), and higher protein levels of the receptors for estrogen, insulin, and IGF-I (all p values ≤0.02). Conversely, pIGF1R/pIR levels were higher in premenopausal proliferative phase versus postmenopausal endometrium (p value =0.01). CONCLUSIONS: These results highlight links between endometrial cancer risk factors and mechanistic factors that may contribute to early events in the multistage process of endometrial carcinogenesis.

Novel epigenetic changes in CDKN2A are associated with progression of cervical intraepithelial neoplasia.

OBJECTIVE: To conduct a comprehensive mapping of the genomic DNA methylation in CDKN2A, which codes for the p16(INK4A) and p14(ARF) proteins, and 14 of the most promising DNA methylation marker candidates previously reported to be associated with progression of low-grade cervical intraepithelial neoplasia (CIN1) to cervical cancer. METHODS: We analyzed DNA methylation in 68 HIV-seropositive and negative women with incident CIN1, CIN2, CIN3 and invasive cervical cancer, assaying 120 CpG dinucleotide sites spanning APC, CDH1, CDH13, CDKN2A, CDKN2B, DAPK1, FHIT, GSTP1, HIC1, MGMT, MLH1, RARB, RASSF1, TERT and TIMP3 using the Illumina Infinium array. Validation was performed using high resolution mapping of the target genes with HELP-tagging for 286 CpGs, followed by fine mapping of candidate genes with targeted bisulfite sequencing. We assessed for statistical differences in DNA methylation levels for each CpG loci assayed using univariate and multivariate methods correcting for multiple comparisons. RESULTS: In our discovery sample set, we identified dose dependent differences in DNA methylation with grade of disease in CDKN2A, APC, MGMT, MLH1 and HIC1, whereas single CpG locus differences between CIN2/3 and cancer groups were seen for CDH13, DAPK1 and TERT. Only those CpGs in the gene body of CDKN2A showed a monotonic increase in methylation between persistent CIN1, CIN2, CIN3 and cancers. CONCLUSION: Our data suggests a novel link between early cervical disease progression and DNA methylation in a region downstream of the CDKN2A transcription start site that may lead to increased p16(INK4A)/p14(ARF) expression prior to development of malignant disease.

MEDULLARY THYROID CANCER THAT STAINS NEGATIVE FOR CA 19-9 HAS DECREASED METASTATIC POTENTIAL.

OBJECTIVE: Presently, no clinical tools are available to diagnose the metastatic potential of medullary thyroid cancer (MTC) at disease presentation. Surveillance with calcitonin (Ct) and carcinoembryonic antigen (CEA) is currently recommended for the observation and diagnosis of metastatic disease after initial treatment of MTC. Recently, carbohydrate antigen (CA)19-9 staining has been associated with aggressive forms of MTC and metastatic spread. This pilot study explored whether positive CA19-9 staining of MTC tissue is associated with its metastatic potential. METHODS: Sixteen cases of MTC were identified, and tissue specimens were immunostained for CA 19-9 and other MTC tumor markers. Clinical information about patients' MTC was collected through a retrospective chart review. RESULTS: Overall, 63% of the specimens stained positive for CA19-9. The median size of positively staining specimens was 2.6 cm (interquartile range [IQR] 1.2-3.2) compared to 0.7 cm (0.5-1.2) in negatively staining MTC specimens (P = .04). All specimens from patients diagnosed with stage IV MTC stained positive for CA19-9, compared to only 40% of cases that were classified as stages I to III (P = .03). Furthermore, 100% of the primary specimens that were documented to have metastatic spread stained positive for CA19-9. The sensitivity for ruling out stage IV MTC based on negative staining for CA 19-9 was 100%. CONCLUSION: Based on these results, we conclude that negative staining of MTC for CA19-9 may be associated with its decreased metastatic potential.

Ecosystem energetic implications of parasite and free-living biomass in three estuaries.

Parasites can have strong impacts but are thought to contribute little biomass to ecosystems. We quantified the biomass of free-living and parasitic species in three estuaries on the Pacific coast of California and Baja California. Here we show that parasites have substantial biomass in these ecosystems. We found that parasite biomass exceeded that of top predators. The biomass of trematodes was particularly high, being comparable to that of the abundant birds, fishes, burrowing shrimps and polychaetes. Trophically transmitted parasites and parasitic castrators subsumed more biomass than did other parasitic functional groups. The extended phenotype biomass controlled by parasitic castrators sometimes exceeded that of their uninfected hosts. The annual production of free-swimming trematode transmission stages was greater than the combined biomass of all quantified parasites and was also greater than bird biomass. This biomass and productivity of parasites implies a profound role for infectious processes in these estuaries.

Selective gene-expression profiling of migratory tumor cells in vivo predicts clinical outcome in breast cancer patients.

INTRODUCTION: Metastasis of breast cancer is the main cause of death in patients. Previous genome-wide studies have identified gene-expression patterns correlated with cancer patient outcome. However, these were derived mostly from whole tissue without respect to cell heterogeneity. In reality, only a small subpopulation of invasive cells inside the primary tumor is responsible for escaping and initiating dissemination and metastasis. When whole tissue is used for molecular profiling, the expression pattern of these cells is masked by the majority of the noninvasive tumor cells. Therefore, little information is available about the crucial early steps of the metastatic cascade: migration, invasion, and entry of tumor cells into the systemic circulation. METHODS: In the past, we developed an in vivo invasion assay that can capture specifically the highly motile tumor cells in the act of migrating inside living tumors. Here, we used this assay in orthotopic xenografts of human MDA-MB-231 breast cancer cells to isolate selectively the migratory cell subpopulation of the primary tumor for gene-expression profiling. In this way, we derived a gene signature specific to breast cancer migration and invasion, which we call the Human Invasion Signature (HIS). RESULTS: Unsupervised analysis of the HIS shows that the most significant upregulated gene networks in the migratory breast tumor cells include genes regulating embryonic and tissue development, cellular movement, and DNA replication and repair. We confirmed that genes involved in these functions are upregulated in the migratory tumor cells with independent biological repeats. We also demonstrate that specific genes are functionally required for in vivo invasion and hematogenous dissemination in MDA-MB-231, as well as in patient-derived breast tumors. Finally, we used statistical analysis to show that the signature can significantly predict risk of breast cancer metastasis in large patient cohorts, independent of well-established prognostic parameters. CONCLUSIONS: Our data provide novel insights into, and reveal previously unknown mediators of, the metastatic steps of invasion and dissemination in human breast tumors in vivo. Because migration and invasion are the early steps of metastatic progression, the novel markers that we identified here might become valuable prognostic tools or therapeutic targets in breast cancer.

Primary bilateral ovarian and uterine Burkitt's lymphoma following chemotherapy for breast cancer.

BACKGROUND: Burkitt's lymphoma is extremely rare in the United States, with reports of approximately 300 new cases each year. A case of Burkitt's lymphoma involving the uterus, cervix, ovaries and appendix 15 years after adjuvant chemotherapy for breast cancer is presented. CASE REPORT: A 58-year-old woman presented with an abdominal mass. MRI revealed a pelvic mass originating from the uterus. Her past medical history was significant for an infiltrating ductal carcinoma of the left breast with positive lymph nodes and extra nodal invasion diagnosed at age 43. She had a left modified radical mastectomy in 1990 with adjuvant chemotherapy. Histologic sections of the uterus, cervix, ovaries, appendix and external iliac lymph node obtained at laparotomy revealed diffuse neoplastic lymphoid infiltration with necrosis. Light microscopy revealed starry sky patterns and immunohistochemical staining demonstrated atypical lymphocytes which stained for CD20, CD10, bcl-6, and Ki-67. After complete staging, chemotherapy was started and the patient is presently tumor free 41 months after the diagnosis. CONCLUSION: Burkitt's lymphoma arising from the uterus/cervix is extremely rare and may present as a pelvic mass. Early diagnosis, aggressive chemotherapy, +/-surgical intervention, plays an important role in management and survival of patients with Burkitt's lymphoma.

Expanding the roles for pregnane X receptor in cancer: proliferation and drug resistance in ovarian cancer.

PURPOSE: We examined the presence of the pregnane X receptor (PXR) and its effects on ovarian cancer cells after activation by its cognate ligand. EXPERIMENTAL DESIGN: SKOV-3 and OVCAR-8 ovarian carcinoma cells were analyzed for expression of PXR by quantitative reverse transcription-PCR and Western blot. Human ovarian cancer tissue was also analyzed for PXR expression by immunochemistry. Ligand (agonist)-induced PXR target genes were analyzed in SKOV-3 cells by quantitative reverse transcription-PCR. SKOV-3 cell proliferation was assessed by MTT assay. In vivo confirmation of in vitro effects of PXR ligands were done in NOD.SCID mice carrying SKOV-3 xenografts. RESULTS: PXR is expressed in ovarian cancer cells. In SKOV-3 cells, PXR is functional and its activation by cognate ligands induces PXR target genes (CYP2B6, CYP3A4, and UGT1A1) but not MDR1 and MRP2. PXR activation in SKOV-3 cells induces cell proliferation and drug resistance. In mice harboring SKOV-3 xenografts, rifampicin (PXR agonist) induces cell proliferation and tumor growth. CONCLUSION: PXR activation, regardless of the type of ligand agonist present, promotes the "malignant" phenotype of cancer cells. These data serve as the basis for finding novel nontoxic inhibitors of PXR activation as a method to control cell growth and prevent induction of drug resistance.

Benign Endocervical Polyp with Heterologous Elements in a 42-year-old Female: Report of a Case.

A 42-year-old female with unremarkable medical history presented for a routine cervical screening upon which an endocervical polyp was identified and submitted entirely for histopathologic evaluation. Microscopic examination showed multiple well-circumscribed nodular fragments of polyp with superficial erosion and focal acute inflammation. Benign endocervical glands were seen within a fibrotic stroma with a prominent smooth muscle component. Intermixed mature adipose tissue and large thick-walled vessels were also identified. Stromal and epithelial atypia were absent; similarly, stromal cellularity, mitoses, and condensation were not identified. Additional deeper levels did not reveal other heterologous elements. The diagnosis of a benign choristomatous endocervical polyp was rendered. As anticipated, the patient recovered completely. A review of the English literature demonstrates rare, namely two other reports of hamartomatous tissue in an endocervical polyp. To the best of our knowledge, we report the third such case.

Cyclin D1 repression of peroxisome proliferator-activated receptor gamma expression and transactivation.

The cyclin D1 gene is overexpressed in human breast cancers and is required for oncogene-induced tumorigenesis. Peroxisome proliferator-activated receptor gamma (PPAR gamma) is a nuclear receptor selectively activated by ligands of the thiazolidinedione class. PPAR gamma induces hepatic steatosis, and liganded PPAR gamma promotes adipocyte differentiation. Herein, cyclin D1 inhibited ligand-induced PPAR gamma function, transactivation, expression, and promoter activity. PPAR gamma transactivation induced by the ligand BRL49653 was inhibited by cyclin D1 through a pRB- and cdk-independent mechanism, requiring a region predicted to form an helix-loop-helix (HLH) structure. The cyclin D1 HLH region was also required for repression of the PPAR gamma ligand-binding domain linked to a heterologous DNA binding domain. Adipocyte differentiation by PPAR gamma-specific ligands (BRL49653, troglitazone) was enhanced in cyclin D1(-/-) fibroblasts and reversed by retroviral expression of cyclin D1. Homozygous deletion of the cyclin D1 gene, enhanced expression by PPAR gamma ligands of PPAR gamma and PPAR gamma-responsive genes, and cyclin D1(-/-) mice exhibit hepatic steatosis. Finally, reduction of cyclin D1 abundance in vivo using ponasterone-inducible cyclin D1 antisense transgenic mice, increased expression of PPAR gamma in vivo. The inhibition of PPAR gamma function by cyclin D1 is a new mechanism of signal transduction cross talk between PPAR gamma ligands and mitogenic signals that induce cyclin D1.

Endangered light-footed clapper rail affects parasite community structure in coastal wetlands.

An extinction necessarily affects community members that have obligate relationships with the extinct species. Indirect or cascading effects can lead to even broader changes at the community or ecosystem level. However, it is not clear whether generalist parasites should be affected by the extinction of one of their hosts. We tested the prediction that loss of a host species could affect the structure of a generalist parasite community by investigating the role of endangered Light-footed Clapper Rails (Rallus longirostris levipes) in structuring trematode communities in four tidal wetlands in southern California, U.S.A. (Carpinteria Salt Marsh, Mugu Lagoon) and Mexico (Estero de Punta Banda, Bahia Falsa-San Quintin). We used larval trematode parasites in first intermediate host snails (Cerithidea californica) as windows into the adult trematodes that parasitize Clapper Rails. Within and among wetlands, we found positive associations between Clapper Rails and four trematode species, particularly in the vegetated marsh habitat where Clapper Rails typically occur. This suggests that further loss of Clapper Rails is likely to affect the abundance of several competitively dominant trematode species in wetlands with California horn snails, with possible indirect effects on the trematode community and changes in the impacts of these parasites on fishes and invertebrates.

Sensitivity of FDG PET, GLUT1 expression and proliferative index in bronchioloalveolar lung cancer.

OBJECTIVE: To estimate the sensitivity of [F] fluorodeoxyglucose (FDG) positron emission tomography (PET) and to assess the expression of glucose transporter 1 (GLUT1) and proliferative index (PI) in bronchioloalveolar lung cancer (BAC). METHODS: Twenty-four patients with resected BAC underwent preoperative PET between October 1996 and February 2003. The surgical specimens were re-examined, and 18 patients who fulfilled the 1999 WHO definition for BAC were included in the study. The PET images were reviewed in order to determine the positive (PET+) and negative (PET-) tumours on PET. The pathology slides were stained with antibodies to GLUT1 and Proliferating cell nuclear antigen (PCNA) in order to determine GLUT1 expression and PI, respectively. RESULTS: There were 13 cases of PET+ BAC (sensitivity, 72%; confidence interval, 52-93%); seven of them were GLUT1+ cases and six were GLUT1-. The stromal cell PI was significantly higher in GLUT1+ BAC compared to GLUT- BAC (50.9+/-17.1 vs. 33.2+/-14.2, P=0.0286), and higher in PET+ BAC compared to PET- BAC (45.5+/-15.3 vs. 29.6+/-19.6, P=0.0854). CONCLUSION: After applying the 1999 WHO criteria, the sensitivity of PET for detecting BAC is still relatively low. Other glucose transporters such as GLUT3 likely play a role in FDG uptake in BAC. GLUT1+ or PET+ BAC tumours have a higher stromal cell PI when compared to GLUT1- BAC or PET- BAC tumours, respectively.

The one that didn't get away.

My part in a lifesaving drama was small... or so I thought.

Preparing for the 21st century: graduate nursing education in Vietnam.

Friendship Bridge Nurses and Vietnamese nurses developed a partnership based on the concepts of collaboration, sustainability, critical mass, and shared vision. This article explores the process of development and issues of implementation of a graduate nursing education program in Vietnam.

Consistency of reporting basic characteristics of lung nodules and masses on computed tomography.

RATIONALE AND OBJECTIVES: To assess the consistency of chest computed tomography (CT) reports in describing basic characteristics of lung nodules and masses. MATERIALS AND METHODS: We retrospectively identified 107 consecutive patients with preoperative chest CT scans before resection of a lung nodule or mass over a 4-year period within a single institution. There were 54 men and 53 women with a mean age of 64 years (range, 37-86) years. The CT scans were reported by a cohort of 20 board-certified radiologists, three of whom reviewed more than 10 CT scans (n = 60 exams). The CT reports were reviewed for lesion characteristics including size, location, and description of margins, presence or absence of calcification, fat and cavitation, and the diagnosis or differential diagnosis. Pathology reports were reviewed for the same characteristics and the final diagnosis. Both CT and pathologic reports of emphysema were noted in lobectomy specimens. The differences between the interpreting radiologists were also sought. RESULTS: A diagnosis or differential diagnosis was provided in 90% (96/107) of CT reports. The diagnosis of bronchogenic carcinoma was made in 78% (59/76) of those with bronchogenic carcinoma, compared with 65% (20/31) of those with other diagnoses (P = NS). Radiologists described the margins of the nodule or mass in 64% (68/107) of cases, similar in frequency to 66% of pathologists (71/107). Radiologic description of an irregular/spiculated margins predicted bronchogenic carcinoma in 86% of cases (42/49), while a smooth/lobulated margins predicted a diagnosis other than bronchogenic carcinoma in 58% (11/19; P < .05). The presence or absence of calcification was noted in 7% (5/76) of cases of bronchogenic carcinoma and 32% (10/31) of those with other diagnoses (P < .05, chi square). Both radiologists and pathologists consistently reported the size of the lesions with a correlation coefficient between radiology and pathology reports of 0.88. CT reporting of the characteristics of the lesion did not differ among lesions of different sizes. There was no significant difference between major reporters (more than 10 cases) in this study. Emphysema in the surrounding lung was reported in 25% (20/81) of radiology and 38% (31/81) of pathology reports (P = NS). CONCLUSION: This series demonstrates a lack of consistent reporting of the margins of resected lung nodules both on CT and on pathologic specimens. The presence or absence of calcification was inconsistently reported, although more frequently noted in diagnoses other than bronchogenic carcinoma. As large-scale CT screening for lung cancer becomes more common, radiologists should prioritize developing and adopting standardized reporting criteria for the CT evaluation of lung nodules.

A clinical and pathologic comparison between stage-matched endometrial intraepithelial carcinoma and uterine serous carcinoma: is there a difference?

Endometrial intraepithelial carcinoma (EIC) is a rare pathologic variant of uterine serous carcinoma (USC). Our aim is to distinguish patterns of clinic-pathologic outcomes in patients with EIC and USC for disease limited to the endometrium (stage 1A) as well as with distant metastasis (stage 4B). Surgically staged patients were retrospectively identified and relevant variables were extracted and compared. Kaplan-Meier was used to generate the survival data. More USC (n = 29) exhibited lymphovascular invasion (stage 4, P = .01) and expressed higher levels of estrogen receptor-α than EIC (P = .0009 and .063 for stages 1 and 4, respectively). The survival is comparable, with 1 recurrence in each group for stage 1A disease. For stage 4 EIC and USC, the progression-free survival (14 vs10 months) and overall survival (19 vs 20 months) are similar to what is previously published. In conclusion, EIC, whether limited to the endometrium, or widely metastatic, imparts similar outcomes and should be treated comparably with stage-matched USC.

Quantification of nucleolar channel systems: uniform presence throughout the upper endometrial cavity.

OBJECTIVE: To determine the prevalence of nucleolar channel systems (NCSs) by uterine region, applying continuous quantification. DESIGN: Prospective clinical study. SETTING: Tertiary care academic medical center. PATIENT(S): Forty-two naturally cycling women who underwent hysterectomy for benign indications. INTERVENTION(S): NCS presence was quantified by a novel method in six uterine regions-fundus, left cornu, right cornu, anterior body, posterior body, and lower uterine segment (LUS)-with the use of indirect immunofluorescence. MAIN OUTCOME MEASURE(S): Percentage of endometrial epithelial cells (EECs) with NCSs per uterine region. RESULT(S): NCS quantification was observer independent (intraclass correlation coefficient 0.96) and its intrasample variability low (coefficient of variation 0.06). Eleven of 42 hysterectomy specimens were midluteal, ten of which were analyzable with nine containing >5% EECs with NCSs in at least one region. The percentage of EECs with NCSs varied significantly between the LUS (6.1%; interquartile range [IQR] 3.0-9.9) and the upper five regions (16.9%; IQR 12.7-23.4), with fewer NCSs in the basal layer of the endometrium (17 ± 6%) versus the middle (46 ± 9%) and luminal layers (38 ± 9%) of all six regions. CONCLUSION(S): NCS quantification during the midluteal phase demonstrates uniform presence throughout the endometrial cavity, excluding the LUS, with a preference for the functional luminal layers. Our quantitative NCS evaluation provides a benchmark for future studies and further supports NCS presence as a potential marker for the window of implantation.

Metastatic medullary thyroid cancer presenting with elevated levels of CA 19-9 and CA 125.

BACKGROUND: Calcitonin and carcinoembryonic antigen (CEA) are established markers of medullary thyroid cancer (MTC), used in the diagnosis and monitoring of disease and its progression. In clinical practice, various other tumor markers are utilized in the follow-up of different malignancies, although their utility has not been well described in MTC. CA 19-9 antigen, routinely used in the monitoring of pancreatic cancer, also has been detected in the tissue of approximately 6% of MTCs. However, its presence has never been reported in the serum of these patients. Elevation of CA 125 antigen, utilized as a tumor marker for ovarian cancer, has never been reported in MTC. We report a novel finding of metastatic MTC presenting with elevated CA 19-9 and CA 125 serum levels, with concurrent tissue staining for these antigens. SUMMARY: A 56-year-old woman with multiple endocrine neoplasia 2B syndrome, post subtotal thyroidectomy for MTC in childhood, presented with extensive metastatic spread of MTC to the lungs and liver, 47 years after the original diagnosis. The patient's calcitonin level decreased from 2950 to 261 pg/mL (reference range: <20 pg/mL) over a 20-year period. The serum CEA level was elevated at 6800 ng/mL (reference range: <5.1 ng/mL). Because of a concern for an alternate malignancy, serum CA 19-9 and CA 125 tumor markers were measured and found to be significantly elevated, at 39,334 U/mL (reference range: <35.1 U/mL) and 96.2 U/mL (reference range: 7-41 U/mL), respectively. Immunostaining of the metastatic MTC tissue showed patchy staining for calcitonin, strongly positive staining for CEA and CA 19-9, and weakly positive staining for CA 125. CONCLUSION: Drawing from experience with CA 19-9 and CA 125 tumor markers in other malignancies, we propose that they may be associated with aggressive forms of MTC with significant metastatic potential.