College of American Pathologists

  Q & A





September 2010

Fredrick L. Kiechle, MD, PhD

Question Q. If a patient sustains a subarachnoid hemorrhage from a ruptured cerebral aneurysm, how long will it take before the appearance of hemosiderin pigment?

A. Because this question has medicolegal implications, we thought it best to discuss the answer at our biannual CAP Neuropathology Committee meeting.

Our consensus is that the very earliest microscopic appearance of hemosiderin pigment within macro-phages would be seen four to five days post-rupture, but the appearance of the pigment would be more obvious seven to 10 days after the aneurysm rupture. Experimental studies on lung alveolar macrophages suggest that very scant hemosiderin pigment can be seen within alveolar macrophages at three days, but again, the peak period is seven to 10 days.

At the other end of the spectrum, the hemosiderin pigment clearance from the leptomeninges is highly variable from one patient to another, in part predicated on patient age and atherosclerosis. Though much of the clearance may be completed by 21 days in most patients, variations in circulatory capacity make it impossible to provide an outside limit for complete clearance. Indeed, in some patients hemosiderin pigmentation of leptomeninges never clears completely, especially after aneurysm rupture.

This may also be because cerebral aneurysms can leak small amounts of blood repeatedly.

Thus, in a patient with a cerebral aneurysm, one is always confronted with the possibility that the aneurysm had been leaking at a low level in the past and that the hemosiderin you are seeing may have been from one of these previous low-level leaks, not from the recent massive rupture.

If this is a question stemming from a medicolegal case in which you are involved, we encourage you to seek primary resources from the literature on this issue.

CAP Neuropathology Committee
Chair, Bette K. DeMasters, MD
University of Colorado

Question Q. Recently I saw a high-grade carcinoma in the hepatic flexure of a 75-year-old man. The tumor was associated with florid Crohn’s-like reaction. MLH1 by immunohistochemistry showed no staining in the tumor cells, but the tumor cells stained for MSH2 and MSH6 proteins by IHC. What is the significance of this isolated finding of loss of MLH1 staining, and should the patient’s children have genetic counseling?

A. The short answer is that the children probably do not need counseling; read on for the long answer.

Loss of MLH1 protein expression by immunohistochemistry is a reliable surrogate marker of high-level microsatellite instability (MSI-H), which occurs in more than 90 percent of cancers related to hereditary nonpolyposis colorectal cancer (HNPCC), or Lynch syndrome.1 MSI-H is also found in 10 percent to 15 percent of sporadic colo-rectal cancers not related to HNPCC.2 HNPCC cancers are caused by mutations in DNA mismatch repair genes, most commonly MLH1 or MSH2, but also MSH6 and PMS2, among others.3 MLH1 mutations account for about 40 percent of HNPCC cancers.3 Determining which MSI-H cancers are related to germline mutations or which are sporadic can require a complex assimilation of all clinicopathologic data, often requiring close collaboration among genetic counselors, gastroenterologists, colorectal surgeons, and pathologists. This is particularly true in cancers demonstrating loss of MLH1 by immunohistochemistry because this is by far the most common phenotype in MSI-H sporadic cancers.2

DNA sequencing of the MLH1 gene remains the gold-standard test in separating germline from sporadic. However, there are several other factors to consider before proceeding to such a labor-intensive and expensive test. Loss of the remaining DNA mismatch repair proteins (MSH2, MSH6, or isolated PMS2) occurs much less commonly in sporadic cancers and warrants strong consideration for germline testing on their own merit. Family and personal cancer history, as described in the Amsterdam II1 or revised Bethesda criteria,3 often serve as the trigger for laboratory-based investigation of HNPCC. Epigenetic MLH1 promoter hypermethylation serves as the most common mechanism of the sporadic MSI-H phenotype,2–4 and testing for MLH1 promoter hypermethylation, or the more global CpG island methylator phenotype (CIMP), can be performed in large reference laboratories on formalin-fixed, paraffinembedded tissue. MLH1 promoter hypermethylation, however, has been shown to occur in HNPCC kindreds2; therefore, this test does not absolutely separate HNPCC cancers from sporadic ones. BRAF V600E mutations, on the other hand, have never been shown to occur in HNPCC cancers associated with MLH1 mutation5; and a BRAF V600E mutation would help to exclude HNPCC. Unfortunately, about 60 percent of sporadic cancers do not show BRAF V600E mutation.5

Regarding the specific patient at hand—our institution’s general policy, as determined by our genetic counselors and colorectal surgeons, is that germline testing is not warranted in patients with MLH1 loss who are older than 71 years and do not have a family history of HNPCC-associated cancers. This determination would be made only after a thorough investigation of the patient’s family history, usually by a genetic counselor.

Aside from the association with HNPCC, MSI-H status also has prognostic and therapeutic importance, although this is somewhat controversial. MSI-H cancers are considered to have improved prognosis stage-for-stage versus their microsatellite stable (MSS) counterparts.6 This prognostic impact occurs despite the relative chemoinsensitivity of MSI-H cancers,6,7 but this lack of beneficial chemotherapeutic effect has not been validated in all trials.8 And, more important, there are very little outcome data in MSI-H cancers with the new standard-of-care treatments that incorporate irinotecan (Folfiri) or oxali-platin (Folfox) into 5-FU-based regimens. Nevertheless, our oncologists occasionally will use the microsatellite instability status of those patients in whom chemotherapy would be of only marginal benefit (for example, stage II cancers) to help guide therapeutic decisions.


1. Vasen HF, Moslein G, Alonso A, et al. Guidelines for the clinical management of Lynch syndrome (hereditary non-polyposis cancer). J Med Genet. 2007;44:353–362.

2. Samowitz WS. Genetic and epigenetic changes in colon cancer. Exp Mol Pathol. 2008;85:64–67.

3. Umar A, Boland CR, Terdiman JP, et al. Revised Bethesda Guidelines for hereditary nonpolyposis colorectal cancer (Lynch syndrome) and microsatellite instability. J Natl Cancer Inst. 2004;96:261–268.

4. Weisenberger DJ, Siegmund KD, Campan M, et al. CpG island methylator phenotype underlies sporadic microsatellite instability and is tightly associated with BRAF mutation in colorectal cancer. Nat Genet. 2006; 38:787–793.

5. Domingo E, Laiho P, Ollikainen M, et al. BRAF screening as a low-cost effective strategy for simplifying HNPCC genetic testing. J Med Genet. 2004;41:664–668.

6. Popat S, Hubner R, Houlston RS. Systematic review of microsatellite instability and colorectal cancer prognosis. J Clin Oncol. 2005;23:609–618.

7. Ribic CM, Sargent DJ, Moore MJ, et al. Tumor microsatellite-instability status as a predictor of benefit from fluorouracil-based adjuvant chemotherapy for colon cancer. N Engl J Med. 2003;349:209–210.

8. Kim GP, Colangelo LH, Wieand HS, et al. Prognostic and predictive roles of high-degree microsatellite instability in colon cancer: a National Cancer Institute–National Surgical Adjuvant Breast and Bowel Project Collaborative Study. J Clin Oncol. 2007;25:754–756.

Thomas Plesec, MD
Assistant Professor of Pathology
Anatomic Pathology
Cleveland Clinic, Cleveland

Dr. Kiechle is medical director of clinical pathology, Memorial Healthcare, Hollywood, Fla.