Diagnostic approach in TFE3-rearranged renal cell carcinoma: a multi-institutional international survey.

Transcription factor E3-rearranged renal cell carcinoma (TFE3-RCC) has heterogenous morphologic and immunohistochemical (IHC) features.131 pathologists with genitourinary expertise were invited in an online survey containing 23 questions assessing their experience on TFE3-RCC diagnostic work-up.Fifty (38%) participants completed the survey. 46 of 50 participants reported multiple patterns, most commonly papillary pattern (almost always 9/46, 19.5%; frequently 29/46, 63%). Large epithelioid cells with abundant cytoplasm were the most encountered cytologic feature, with either clear (almost always 10/50, 20%; frequently 34/50, 68%) or eosinophilic (almost always 4/49, 8%; frequently 28/49, 57%) cytology. Strong (3+) or diffuse (>75% of tumour cells) nuclear TFE3 IHC expression was considered diagnostic by 13/46 (28%) and 12/47 (26%) participants, respectively. Main TFE3 IHC issues were the low specificity (16/42, 38%), unreliable staining performance (15/42, 36%) and background staining (12/42, 29%). Most preferred IHC assays other than TFE3, cathepsin K and pancytokeratin were melan A (44/50, 88%), HMB45 (43/50, 86%), carbonic anhydrase IX (41/50, 82%) and CK7 (32/50, 64%). Cut-off for positive TFE3 fluorescent in situ hybridisation (FISH) was preferably 10% (9/50, 18%), although significant variation in cut-off values was present. 23/48 (48%) participants required TFE3 FISH testing to confirm TFE3-RCC regardless of the histomorphologic and IHC assessment. 28/50 (56%) participants would request additional molecular studies other than FISH assay in selected cases, whereas 3/50 participants use additional molecular cases in all cases when TFE3-RCC is in the differential.Optimal diagnostic approach on TFE3-RCC is impacted by IHC and/or FISH assay preferences as well as their conflicting interpretation methods.

Incidence and distribution of UroSEEK gene panel in a multi-institutional cohort of bladder urothelial carcinoma.

Noninvasive approaches for early detection of bladder cancer are actively being investigated. We recently developed a urine- based molecular assay for the detection and surveillance of bladder neoplasms (UroSEEK). UroSEEK is designed to detect alterations in 11 genes that include most common genetic alterations in bladder cancer. In this study, we analyzed 527 cases, including 373 noninvasive and 154 invasive urothelial carcinomas of bladder from transurethral resections or cystectomies performed at four institutions (1991-2016). Two different mutational analysis assays of a representative tumor area were performed: first, a singleplex PCR assay for evaluation of the TERT promoter region (TERTSeqS) and second, a multiplex PCR assay using primers designed to amplify regions of interest of 10 (FGFR3, PIK3CA, TP53, HRAS, KRAS, ERBB2, CDKN2A, MET, MLL, and VHL) genes (UroSeqS). Overall, 92% of all bladder tumors were positive for at least one genetic alteration in the UroSEEK panel. We found TERT promoter mutations in 77% of low-grade noninvasive papillary carcinomas, with a relatively lower incidence of 65% in high-grade noninvasive papillary carcinomas and carcinomas in situ; p = 0.017. Seventy-two percent of pT1 and 63% of muscle-invasive bladder tumors harbored TERT promoter mutations with g.1295228C>T alteration being the most common in all groups. FGFR3 and PIK3CA mutations were more frequent in low-grade noninvasive papillary carcinomas compared with high-grade noninvasive papillary carcinomas and carcinomas in situ (p < 0.0001), while the opposite was true for TP53 (p < 0.0001). Significantly higher rates of TP53 and CDKN2A mutation rates (p = 0.005 and 0.035, respectively) were encountered in muscle-invasive bladder tumors compared with those of pT1 stage. The overwhelming majority of all investigated tumors showed at least one mutation among UroSEEK assay genes, confirming the comprehensive coverage of the panel and supporting its potential utility as a noninvasive urine-based assay.

Targeted sequencing of plasmacytoid urothelial carcinoma reveals frequent TERT promoter mutations.

Activating mutations in the promoter of the telomerase reverse transcriptase (TERT) gene are the most common genetic alterations in urothelial carcinoma (UC) of the bladder and upper urinary tract. Although the cadherin 1 (CDH1) gene is commonly mutated in the clinically aggressive plasmacytoid variant of urothelial carcinoma (PUC), little is known about their TERT promoter mutation status. A retrospective search of our archives for PUC and UC with plasmacytoid and/or signet ring cell features (2007-2014) was performed. Ten specimens from 10 patients had archived material available for DNA analysis and were included in the study. Intratumoral areas of nonplasmacytoid histology were also evaluated when present. Samples were analyzed for TERT promoter mutations with Safe-SeqS, a sequencing error-reduction technology, and sequenced using a targeted panel of the 10 most commonly mutated genes in bladder cancer on the Illumina MiSeq platform. TERT promoter mutations were detected in specimens with pure and focal plasmacytoid features (6/10). Similar to conventional UC, the predominant mutation identified was g.1295228C>T. In heterogeneous tumors with focal variant histology, concordant mutations were found in plasmacytoid and corresponding conventional, glandular, or sarcomatoid areas. Co-occurring mutations in tumor protein p53 (TP53, 2 cases) and kirsten rat sarcoma (KRAS) viral proto-oncogene (1 case) were also detected. TERT promoter mutations are frequently present in PUC, which provides further evidence that TERT promoter mutations are common events in bladder cancer, regardless of histologic subtype, and supports their inclusion in any liquid biopsy assay for bladder cancer.

Successful Treatment of Posttransplant Recurrent Complement C3 Glomerulopathy with Eculizumab.

Two-thirds of complement C3 glomerulopathy (C3G) recur after transplantation and commonly cause graft loss. There is not a standard treatment protocol for these cases. We present a kidney transplant patient with recurrent C3G who was successfully treated with eculizumab. Nephrotic proteinuria and hematuria occurred and creatinine levels increased after transplantation. A graft biopsy revealed recurrent C3G. The patient was administered 250 mg pulse methylprednisolone for 3 days and had 9 sessions of plasmapheresis. Since elevated creatinine levels and proteinuria persisted, eculizumab was instituted. A complete remission was observed after 9-month maintenance eculizumab treatment. Eculizumab may be a potentially effective option in kidney transplant patients with recurrent C3G unresponsive to other treatment modalities.

Non-invasive detection of urothelial cancer through the analysis of driver gene mutations and aneuploidy.

Current non-invasive approaches for detection of urothelial cancers are suboptimal. We developed a test to detect urothelial neoplasms using DNA recovered from cells shed into urine. UroSEEK incorporates massive parallel sequencing assays for mutations in 11 genes and copy number changes on 39 chromosome arms. In 570 patients at risk for bladder cancer (BC), UroSEEK was positive in 83% of those who developed BC. Combined with cytology, UroSEEK detected 95% of patients who developed BC. Of 56 patients with upper tract urothelial cancer, 75% tested positive by UroSEEK, including 79% of those with non-invasive tumors. UroSEEK detected genetic abnormalities in 68% of urines obtained from BC patients under surveillance who demonstrated clinical evidence of recurrence. The advantages of UroSEEK over cytology were evident in low-grade BCs; UroSEEK detected 67% of cases whereas cytology detected none. These results establish the foundation for a new non-invasive approach for detection of urothelial cancer.

Spectrum of genetic mutations in de novo PUNLMP of the urinary bladder.

Our group and others have previously demonstrated the presence of TERT promoter mutations (TERT-mut) in 60-80% of urothelial carcinomas and some of their histologic variants. Five other genes have been frequently implicated in bladder cancer: FGRF3, TP53, PIK3CA, HRAS, and CDKN2A. In the current study, we sought to determine the prevalence of mutations in TERT and these five other genes in de novo papillary urothelial neoplasms of low malignant potential (PUNLMP) of the urinary bladder. A retrospective search of our archives for PUNLMP was performed and 30 de novo cases were identified and included in the study. We found mutations in TERT (TERT-mut) and FGFR3 (FGFR3-mut) to be the most common alterations in the cohort (63 and 60%, respectively). The majority of the TERT-mut-positive tumors (84%) had a g.1295228C > T alteration with the remaining tumors demonstrating g.1295250C > T. Approximately one fourth of tumors had TP53 mutations. These findings support the potential utility of a uniform genetic mutation panel to detect bladder cancers of various subtypes.

High prevalence of TERT promoter mutations in micropapillary urothelial carcinoma.

Somatic activating mutations in the promoter of the telomerase reverse transcriptase (TERT) gene are the most common genetic alterations in urothelial carcinoma (UC) of the bladder and upper urinary tract. Little is known, however, about TERT-mutation status in the relatively uncommon but clinically aggressive micropapillary (MPC) variant. We evaluated the presence of TERT promoter mutations in MPC of the bladder and upper urinary tract. A retrospective search of our archives for MPC and UC with micropapillary features (2005-2014) was performed. All slides were reviewed to confirm the histologic diagnosis. Thirty-three specimens from 31 patients had FFPE blocks available for DNA analysis and were included in the study. Intratumoral areas of non-micropapillary histology were also evaluated when present. Samples were analyzed with Safe-SeqS, a sequencing error reduction technology, and sequenced using the Illumina MiSeq platform. TERT promoter mutations were detected in all specimens with pure MPC (18 of 18) and UC with focal micropapillary features (15 of 15). Similar to conventional UC, the predominant mutations identified occurred at positions -124 (C228T) (85 %) and -146 (C250T) (12 %) bp upstream of the TERT ATG start site. In heterogeneous tumors with focal variant histology, intratumoral concordant mutations were found in variant (MPC and non-MPC) and corresponding conventional UC. We found TERT promoter mutations, commonly found in conventional UC, to be frequently present in MPC. Our finding of concordant intratumoral mutational alterations in cases with focal variant histology lends support to the common oncogenesis origin of UC and its variant histology.

High prevalence of TERT promoter mutations in primary squamous cell carcinoma of the urinary bladder.

TERT promoter mutations (TERT-mut) are detectable in the majority of urothelial carcinomas. The detection of TERT-mut in urine is under investigation as a potential urine-based molecular-screening assay for bladder cancer. A small but significant number of bladder carcinomas are pure squamous cell carcinoma. We sought to assess the incidence of TERT-mut in squamous cell carcinoma of the urinary bladder. A retrospective search of the institutional pathology archives yielded 15 cystectomy specimens performed for squamous cell carcinoma (2000-2014). Histologic slides were reviewed by a senior urologic pathologist to confirm the diagnosis and select a representative formalin-fixed paraffin-embedded tissue block for mutational analysis. All cases yielded adequate material for DNA analysis. Sequencing for TERT-mut was performed using previously described SafeSeq technique. We detected TERT-mut in 12/15 (80%) of bladder squamous cell carcinomas. TERT promoter mutations, commonly found in conventional urothelial carcinoma, are also highly prevalent in urinary bladder squamous cell carcinoma suggesting a common tumorigenesis and potential utility as a molecular urine-based-screening assay.

Detection of TERT promoter mutations in primary adenocarcinoma of the urinary bladder.

TERT promoter mutations (TERT-mut) have been detected in 60% to 80% of urothelial carcinomas. A molecular urine-based screening assay for the detection of TERT-mut is currently being pursued by our group and others. A small but significant number of bladder carcinomas are adenocarcinoma. The current study assesses the incidence of TERT-mut in primary adenocarcinomas of urinary bladder. A retrospective search of our institutional pathology records identified 23 cystectomy specimens with a diagnosis of adenocarcinoma (2000-2014). All slides were reviewed by a senior urologic pathologist to confirm tumor type and select a representative formalin-fixed, paraffin-embedded block for mutational analysis. Adequate material for DNA testing was available in 14 cases (7 enteric type and 7 not otherwise specified). TERT-mut sequencing analysis was performed using previously described SafeSeq technique. Overall, 28.5% of primary adenocarcinoma harbored TERT-mut. Interestingly, 57% of nonenteric adenocarcinomas were mutation positive, whereas none of the enteric-type tumors harbored mutations. Similar to urothelial carcinoma, we found a relatively higher rate of TERT-mut among nonenteric-type adenocarcinomas further supporting the potential utility of TERT-mut urine-based screening assay for bladder cancer.

The role of GATA binding protein 3 in the differential diagnosis of collecting duct and upper tract urothelial carcinomas.

Differential diagnosis of collecting duct carcinoma (CDC) from invasive upper tract urothelial carcinoma (UTUC) can be challenging. PAX8 and p63 are 2 markers often used in this setting. GATA binding protein 3 (GATA3) is a marker of urothelial differentiation. We investigated GATA3 expression in CDC and UTUC and its use in this differential. Eighteen CDC and 25 UTUC cases were used to build 2 tissue microarrays. GATA3, p63, and PAX8 nuclear expression was evaluated using standard immunohistochemistry. Staining intensity and percentage of positive cells were assessed. Sensitivity, specificity, and positive and negative predictive values of the markers and their combination were also evaluated. We found GATA3 positivity in 22 (88%) of 25 UTUCs and 1 (6%) of 18 CDCs. The median GATA3 extent of expression was higher in UTUC than in CDC (74% versus 0%, P = .00). We found p63 positivity in 23 (92%) of 25 UTUCs and 2 (11%) of 18 CDCs. PAX8 was positive in 3 (12%) of 25 UTUCs and all (100%) CDCs. GATA3 sensitivity and specificity for UTUC were 88% and 94%, respectively. p63 sensitivity and specificity for UTUC were 92% and 89%, respectively. The p63+/PAX8- profile showed higher sensitivity for UTUC than did the GATA3+/PAX8- profile (80% versus 76%). Both showed a specificity of 100% for UTUC. GATA3+ or p63+/PAX8- sensitivity and specificity for UTUC were 84% and 100%, respectively. Immunohistochemical expression of GATA3 was higher in UTUC, suggesting a potential role for distinguishing UTUC from CDC. Adding this marker to the combination panel of p63 and PAX8 might improve its performance in the diagnosis of epithelial neoplasms involving the renal sinus.

PAX8 (+)/p63 (-) immunostaining pattern in renal collecting duct carcinoma (CDC): a useful immunoprofile in the differential diagnosis of CDC versus urothelial carcinoma of upper urinary tract.

BACKGROUND: Collecting duct carcinoma (CDC) is a relatively rare but aggressive type of renal malignancy with variable morphologic features. One of the World Health Organization diagnostic criteria for CDC is the exclusion of urothelial carcinoma of renal pelvis from the differential diagnosis. PAX8 is a novel lineage restricted transcription factor expressed in renal tubules. We investigated the expression pattern of PAX8 in CDC and its utility, in combination with p63, in resolving the differential diagnosis of CDC versus upper tract urothelial carcinoma (UUC). DESIGN: Archival tissues from 21 CDC and 34 UUC were retrieved from our institutional files. Immunohistochemistry for PAX8 and p63 were performed on routine and tissue microarray sections using standard immunohistochemistry protocol. Intensity of nuclear staining was evaluated for each marker and assigned an incremental 0, 1+, 2+, and 3+ score. Extent of staining was categorized as focal (<25%), nonfocal (25% to 75%), or diffuse (>75%). RESULTS: CDC: All 21 (100%) CDC were positive for PAX8. Intensity of expression was moderate to strong (2+/3+) in 19 cases (90%). Extent of staining was diffuse in 13 of 21 tumors. The p63 was positive in 3 of 21 (14%) CDC cases (PAX8+/p63+). UUC: The 34 UUC included 5 pT1, 4 pT2, and 25 pT3/pT4 tumors. Thirty-one of 34 (91.2%) UUC were negative for PAX8, whereas 33 of 34 (97%) were p63 positive. Staining intensity was moderate in 15 cases (44%), of which 12 were nonfocal or diffuse. The unique p63-negative UUC was a pT1 tumor that was also negative for PAX8 (PAX8-/p63-). CONCLUSIONS: We propose the use of the combination of PAX8 and p63 in the diagnosis of poorly differentiated renal sinus epithelial neoplasms where the differential diagnosis includes CDC versus UUC. The immunoprofile of PAX8+/p63- supports the diagnosis of CDC with a sensitivity of 85.7% and a specificity of 100%. In contrast, a (PAX8-/p63+) profile supports the diagnosis of UUC with a sensitivity of 88.2% and a specificity of 100%. The inverse PAX8/p63 expression seen in CDC and UUC supports a renal tubular rather than an urothelial differentiation in CDC given the nephric lineage restriction of PAX8.

Renal cell carcinoma with inferior vena cava thrombus: the Hacettepe experience.

OBJECTIVES: We evaluated the clinical outcome and factors affecting survival in patients with renal cell carcinoma (RCC) and tumor thrombus involving inferior vena cava (IVC). METHODS: Between 1990 and 2007, 28 patients with RCC and tumor thrombus extending into IVC underwent radical nephrectomy and thrombectomy. Patient data were reviewed retrospectively to evaluate the demographics, clinical presentation, surgical approach, pathological features, clinical outcomes, and survival. RESULTS: Twenty-eight patients with a mean age of 52.7 years were operated. Thrombus level was infrahepatic in 15 patients (54%), intrahepatic in 3 patients (10%), suprahepatic in 3 patients (10%), supradiaphragmatic in 2 patients (8%), and intracardiac in 5 patients (18%). All patients with intracardiac thrombi underwent cardiopulmonary bypass (CPB) and deep hypothermic circulatory arrest (DHCA). The mean tumor size was 98.21 mm. Four patients had distant metastases and 3 patients had lymph node involvement. Pathological examination revealed RCC of clear cell type in 26 patients, papillary in 1 and chromophobe in 1 patient. At a mean follow-up of 36.4 months, 16 patients were still alive while 8 patients died due to disease progression and 2 patients died of other causes. Two patients died of pulmonary emboli in the early postoperative period. Lymph node involvement, distant metastases, hypercalcemia, and sarcomatoid component were found to be factors affecting overall survival significantly. Level of tumor thrombus and Fuhrman grade did not affect survival. CONCLUSIONS: Radical nephrectomy and tumor thrombectomy is currently known to be the most effective method in patients with RCC and tumor thrombus extending into IVC. Factors affecting survival are the ones related to tumor biology. Tumor thrombus level does not affect the prognosis.

Effects of amifostine on glycerol-pretreated rabbit kidneys.

Glycerol-induced acute renal failure is an experimental model for myoglobinuric nephropathy. Amifostine is a cytoprotective agent which scavenges the free radicals. Since there is enhanced production of reactive oxygen metabolites in glycerol-induced acute renal failure, we wanted to examine whether amifostine has a protective role against vascular reactivity and histological changes in kidneys isolated from glycerol-pretreated rabbits. Perfusion pressure was recorded from kidneys obtained from rabbits injected with glycerol 3 hr before the experiments and from glycerol-pretreated and non-pretreated rabbits injected with amifostine 30 min. before the experiments. Acetylcholine-induced (10(-8)-10(-5) M) vasodilatation was tested following the construction of submaximal vasoconstriction by phenylephrine. Histological investigation was performed using light microscope. Acetylcholine-induced vasodilatation was found to be significantly decreased in glycerol, glycerol+amifostine and amifostine groups compared to controls at all concentrations. Reduction in acetylcholine-induced vasodilation was more prominent in amifostine group compared to amifostine+glycerol group. There was histological renal damage in all experimental groups and this damage was more pronounced in glycerol+amifostine group. In conclusion, contrary to expectation, amifostine per se led to histological damage and potentiated the histological damage caused by glycerol and produced a decrease in acetylcholine-induced vasodilatation. The mechanisms by which amifostine exerts its effects are not known.

Preventive effect of pentoxifylline on renal scarring in rat model of pyelonephritis.

OBJECTIVES: To evaluate the efficiency of pentoxifylline (PTX), a methyl xanthine derivative, in preventing renal scar formation after the induction of pyelonephritis in an experimental rat model with delayed antimicrobial therapy. METHODS: An inoculum of 1 x 10(9) colony-forming units/0.1 mL of the K-12 strain of Escherichia coli, which has both type 1 and P pili, was injected directly into both renal parenchyma of Wistar rats (n = 40). Group 1 (control) received isotonic saline instead of bacterial solution (n = 10). Four equal groups were then formed: group 2 was not treated and group 3 was treated only with ciprofloxacin for 5 days, starting 3 days after bacterial inoculation; in group 4, 50 mg/kg of PTX, and in group 5, PTX (50 mg/kg) and ciprofloxacin (15 mg/kg) together were administered intraperitoneally for 5 days, starting 3 days after bacterial inoculation. Six weeks after bacterial inoculation, all the rats were killed, and both kidneys were examined histopathologically for renal scarring. RESULTS: Delayed treatment with antibiotics had no effect on scarring compared with the untreated controls. However, the addition of PTX to the delayed antibiotic therapy significantly inhibited renal scarring compared with the untreated or antibiotic-only groups (P <0.05). CONCLUSIONS: These results suggest that PTX is effective in preventing renal scar formation in pyelonephritis when the initiation of antimicrobial treatment is delayed in this rat model of pyelonephritis.

Giant eccrine acrospiroma: A case report and review of the literature.

BACKGROUND: Acrospiromas are histologically distinct cutaneous tumours of sweat duct origin and usually measure 1 to 2 cm in size. OBJECTIVE: We describe a patient with a large benign eccrine acrospiroma. METHODS: Case report and literature review. RESULTS: A 50-year-old woman underwent excision of a 5.5×5.0 cm cystic mass located on the left gluteal region, and the resulting defect was closed by a local flap. A diagnosis of benign eccrine acrospiroma was made after histopathological examination. On histological examination, the lesion was characterized by multilobular dermal masses composed of a biphasic cell population. There was no recurrence five months after the operation. CONCLUSION: Although eccrine acrospiromas are usually benign, they can, on rare occasions, undergo malignant transformation. In addition, the clinical appearance of this lesion is not specific and differential diagnosis from other lesions, both benign and malignant, can only be done after the complete removal of the lesion.

RENSEIGNEMENTS GÉNÉRAUX: Les acrospiromes sont des tumeurs cutanées en rapport avec le conduit sudorifère, histologiquement distinctes et mesurant habituellement entre 1 et 2 cm. OBJECTIF: Nous décrivons le cas d'une patiente présentant une acrospirome eccrine bénigne étendue. MÉTHODOLOGIE: Observation de cas et survol de la littérature. RÉSULTATS: Une femme de 50 ans a subi l'ablation d'une masse kystique de 4,5'4,0 cm siégeant dans la région fessière; le vide occasionné par l'ablation du kyste a été comblé au moyen d'un lambeau local. Un diagnostic d'acrospirome eccrine bénigne a été posé après examen histologique. Cet examen a également permis de préciser que la lésion était formée d'une masse dermique multilobulée composée d'une population cellulaire biphasique. On n'a pas observé de récidive cinq mois après l'opération. CONCLUSION: Bien que les acrospiromes eccrines soient habituellement bénignes, elles peuvent, quoique rarement, subir une transformation maligne. En outre, il s'agit d'une lésion ne présentant aucun caractère clinique spécifique de sorte que la distinction avec d'autres lésions, bénignes ou malignes, ne peut être faite qu'après excision complète de la lésion.

Effects of growth factors on doxorubicin-induced skin necrosis: documentation of histomorphological alterations and early treatment by GM-CSF and G-CSF.

Extravasation of vesicant antineoplastic agents such as doxorubicin into the skin or subcutaneous tissues may result in loss of the full thickness of the skin or underlying structures. Several treatment methods have been advocated but none has demonstrated any superiority to the others. The authors designed a controlled animal study in 88 rats to test three methods of early treatment of extravasation of the vesicant antineoplastic agent doxorubicin. The first step of the study included 48 Sprague-Dawley rats. All animals received intradermal injections of 1 mg doxorubicin superficially to the panniculus carnosus in the dorsum. The rats were then divided into four groups of 12 rats each, as follows: group 1, no treatment; group 2, immediate intradermal injection of 0.1 ml saline to the same site; group 3, immediate intradermal injection of 10 microg granulocyte macrophage-colony stimulating factor (GM-CSF) in 0.1 ml saline to the same site; group 4, immediate intradermal injection of 10 microg granulocyte-colony stimulating factor (G-CSF) in 0.1 ml saline to the same site. During the next 6 weeks the rats were observed for the development of necrosis. Ulcers developed and reached maximum size two weeks after the injections. The largest ulcers according to area were observed in group 1 and the mean value was 21.25 mm (p < 0.05). Although wound areas were significantly smaller in the saline group than in the control group and the mean value was 7.58 mm (p < 0.05), the smallest lesions were observed in groups 3 and 4, and the mean values were 1.08 mm and 0.83 mm respectively (p < 0.05). There was statistically no difference with regard to mean ulcer area between groups 3 and 4. During the second step of the experiment, the remaining 40 Sprague-Dawley rats were used. Groups containing 10 rats each were designed similarly after all animals received intradermal injections of 1 mg doxorubicin into the back. On the 10th day after the injection, the entire area of the ulcer together with the underlying panniculus carnosus was excised for pathological examination and for determination of glucose 6-phosphate dehydrogenase (G6PD) activity. On microscopic examination, the extravasated ulcer consisted of a large area of ischemic necrosis. There was marked damage to small blood vessels in the form of fibrinoid necrosis and vasculitis. Injured vessel counts were higher in the control group (group 1; p < 0.05). No difference was observed in G6PD activity between the groups. The authors conclude that both saline and tissue growth factors (GM-CSF and G-CSF) are useful for the early treatment of doxorubicin extravasation; however, GM-CSF and G-CSF are more beneficial.