[The Iceman : Life scenarios and pathological findings from 30 years of research on the glacier mummy "Ötzi"]. / Der Mann aus dem Eis : Lebensszenario und Pathologische Befunde aus 30 Jahren Forschung an der Gletschermumie "Ötzi".

The comprehensive investigation of the excellently preserved mummy of Ötzi, the Iceman, and his equipment over the last 30 years has provided a wealth of information about the life and disease of this late Neolithic individual. This research has indicated that his origin was from a local southern Alpine population, that he grew up in the valleys of the Southern Alps, and that he had considerable local mobility. He had well-balanced nutrition with a mixed vegetable and animal diet. He was very mobile in the alpine terrain and of athletic constitution. The Iceman suffered from mild to moderate degenerative joint disease primarily of the right hip joint, slight spondylosis of the cervical and lumbar spine, a minor focal (premature) arteriosclerosis, lung anthracosis and possibly silicosis, previous pleuritic inflammation (possibly of post-specific origin), intestinal infections of the stomach by Helicobacter pylori and Trichuris trichiura worm infestation in the intestines, a mild osteomalacia of cancellous bone, and diverse pathologies of his teeth with dental caries and periodontitis, as well as hair anomalies. The presence of borreliosis is still under debate. As potential remedies, the Iceman carried some anthelmintic substances with him: a birch polypore and an anthelmintic fern. The numerous tattoos may also have had therapeutic effects. Finally, the last days of Ötzi could be reconstructed quite precisely: his gastrointestinal content indicates that the Iceman moved from Alpine heights to a lower location and then again up to the glacier region where he died. During this journey he encountered two attacks: the first, several days before his death, lead to a stabbing wound in his right hand; the second was an arrow hit that wounded the Iceman lethally at his left axilla by laceration of the subclavian artery.

Paleoproteomic study of the Iceman's brain tissue.

The Tyrolean Iceman, a Copper-age ice mummy, is one of the best-studied human individuals. While the genome of the Iceman has largely been decoded, tissue-specific proteomes have not yet been investigated. We studied the proteome of two distinct brain samples using gel-based and liquid chromatography-mass spectrometry-based proteomics technologies together with a multiple-databases and -search algorithms-driven data-analysis approach. Thereby, we identified a total of 502 different proteins. Of these, 41 proteins are known to be highly abundant in brain tissue and 9 are even specifically expressed in the brain. Furthermore, we found 10 proteins related to blood and coagulation. An enrichment analysis revealed a significant accumulation of proteins related to stress response and wound healing. Together with atomic force microscope scans, indicating clustered blood cells, our data reopens former discussions about a possible injury of the Iceman's head near the site where the tissue samples have been extracted.

Usefulness of p16ink4a, ProEX C, and Ki-67 for the diagnosis of glandular dysplasia and adenocarcinoma of the cervix uteri.

Although the diagnostic criteria of in-situ and invasive adenocarcinomas of the cervix uteri are well established, the differentiation from benign mimics may be difficult and the morphologic features of the precursors of endocervical adenocarcinoma are still debated. In this study, we evaluated the usefulness of p16ink4a (p16), ProEX C, and Ki-67 for the diagnosis of endocervical adenocarcinoma and its precursors. Immunohistochemistry with p16, ProEX C, and Ki-67 was performed in 82 glandular lesions including 15 invasive adenocarcinomas, 29 adenocarcinomas in situ (AIS), 22 non-neoplastic samples, and 16 cases of glandular dysplasia (GD), which showed significant nuclear abnormalities but did not meet the diagnostic criteria for AIS. The immunohistochemical expression pattern was scored according to the percentage of the stained cells (0, 1+, 2+, and 3+ when 0% to 5%, 6% to 25%, 26% to 50%, and more than 50% of the cells were stained, respectively) and was evaluated for each antibody. p16 was at least focally expressed (1+ or more) in 14 of 15 invasive adenocarcinomas, in all AIS and in 7 negative samples. ProEX C and Ki-67 both scored 1+ or more in all adenocarcinomas and AIS and in 8 and 6 negative samples, respectively. Of the GD 15, 14, and 15 expressed p16, ProEX C, and Ki-67, respectively. The score differences between neoplastic and non-neoplastic samples were highly significant for each marker (P<0.001); however, the score distribution by marker differed significantly only in GD (P=0.006) in which, compared with the other markers, p16 showed more often a 3+ pattern. Our study shows that p16, Ki-67, and ProEX C may be helpful for the diagnosis of glandular lesions of the cervix uteri and may also improve the diagnostic accuracy of endocervical GD. In particularly problematic cases, the combination of p16 and a proliferation marker can provide additional help for the interpretation of these lesions.

Genotyping human ancient mtDNA control and coding region polymorphisms with a multiplexed Single-Base-Extension assay: the singular maternal history of the Tyrolean Iceman.

BACKGROUND: Progress in the field of human ancient DNA studies has been severely restricted due to the myriad sources of potential contamination, and because of the pronounced difficulty in identifying authentic results. Improving the robustness of human aDNA results is a necessary pre-requisite to vigorously testing hypotheses about human evolution in Europe, including possible admixture with Neanderthals. This study approaches the problem of distinguishing between authentic and contaminating sequences from common European mtDNA haplogroups by applying a multiplexed Single-Base-Extension assay, containing both control and coding region sites, to DNA extracted from the Tyrolean Iceman. RESULTS: The multiplex assay developed for this study was able to confirm that the Iceman's mtDNA belongs to a new European mtDNA clade with a very limited distribution amongst modern data sets. Controlled contamination experiments show that the correct results are returned by the multiplex assay even in the presence of substantial amounts of exogenous DNA. The overall level of discrimination achieved by targeting both control and coding region polymorphisms in a single reaction provides a methodology capable of dealing with most cases of homoplasy prevalent in European haplogroups. CONCLUSION: The new genotyping results for the Iceman confirm the extreme fallibility of human aDNA studies in general, even when authenticated by independent replication. The sensitivity and accuracy of the multiplex Single-Base-Extension methodology forms part of an emerging suite of alternative techniques for the accurate retrieval of ancient DNA sequences from both anatomically modern humans and Neanderthals. The contamination of laboratories remains a pressing concern in aDNA studies, both in the pre and post-PCR environments, and the adoption of a forensic style assessment of a priori risks would significantly improve the credibility of results.

The Iceman's Last Meal Consisted of Fat, Wild Meat, and Cereals.

The history of humankind is marked by the constant adoption of new dietary habits affecting human physiology, metabolism, and even the development of nutrition-related disorders. Despite clear archaeological evidence for the shift from hunter-gatherer lifestyle to agriculture in Neolithic Europe [1], very little information exists on the daily dietary habits of our ancestors. By undertaking a complementary -omics approach combined with microscopy, we analyzed the stomach content of the Iceman, a 5,300-year-old European glacier mummy [2, 3]. He seems to have had a remarkably high proportion of fat in his diet, supplemented with fresh or dried wild meat, cereals, and traces of toxic bracken. Our multipronged approach provides unprecedented analytical depth, deciphering the nutritional habit, meal composition, and food-processing methods of this Copper Age individual.

Laminin-5 gamma2 chain immunohistochemistry facilitates the assessment of invasiveness and improves the diagnostic reproducibility of glandular lesions of the cervix uteri.

The aim of this study was to evaluate the influence of laminin-5 (LN-5) gamma2 chain immunohistochemistry on the assessment of invasiveness in cervical adenocarcinomas and its impact on the diagnostic reproducibility of glandular lesions of the cervix uteri. Immunohistochemistry with LN-5 gamma2 was performed on 30 cases, including 12 adenocarcinomas in situ (AISs), 5 AISs that were suggestive, albeit not conclusive, of infiltration (AIS+), 7 frankly invasive adenocarcinomas, and 6 nonneoplastic cases with reactive changes. Diagnostic agreement between 3 observers was evaluated by kappa statistics in routine histologic specimens and with the aid of LN-5 gamma2 immunohistochemistry. Laminin-5 gamma2 was expressed in 5 of the 12 AISs (41.6%), all AIS+ and invasive adenocarcinomas, and none of the reactive cases. Cytoplasmatic staining was detected at the invasion front of frankly invasive adenocarcinomas and in tumor buds of all AISs with minimal stromal infiltration. Overall, interobserver agreement was significantly improved by adding LN-5 gamma2 immunostains to the conventional slides (0.56 versus 0.86; P = .002). The difference in interobserver agreement further increased when including only AISs and AIS+ in the analysis (0.17 versus 0.72; P = .000). After immunohistochemical evaluation, the original AIS diagnosis was unanimously changed to adenocarcinoma with minimal stromal invasion in 3 of 12 cases (25%), whereas a discordant hematoxylin-eosin diagnosis turned into a concordant one in 10 of 13 cases (6 AISs, 2 AIS+, 2 adenocarcinomas; 76.9%). Immunohistochemistry with LN-5 gamma2 facilitates the assessment of the invasiveness of cervical adenocarcinomas and improves the interobserver agreement in glandular lesions of the cervix uteri.

Paraneoplastic stiff-person syndrome, heterotopic soft tissue ossification and gonarthritis in a HLA B27-positive woman preceding the diagnosis of Hodgkin's lymphoma.

Paraneoplastic neurologic syndromes associated with Hodgkin's lymphoma include the stiff-person syndrome. A case of stiff-person syndrome is reported who first presented with muscular hyperactivity and acute respiratory failure followed by heterotopic soft tissue ossification and acute seronegative gonarthitis. Initial improvement of a tetanus-like clinical picture was achieved with benzodiazepam given by continuous infusion for analgo-sedation to mechanically ventilate the patient followed by baclofen after successful weaning. The patient was HLA B27 positive and on conventional testing no autoantibodies were detected including anti-glutamic acid decarboxylase antibodies (anti-GAD). Months later in the absence of signs of stiff-person syndrome, mediastinal lymphadenopathy and pleural effusions developed which were diagnosed as classical Hodgkin's lymphoma that was successfully treated with polychemotherapy. No relapse of paraneoplastic neurologic syndromes was seen after two years of lymphoma remission. The case illustrates that stiff-person syndrome may precede the clinical appearance of symptomatic Hodgkin's lymphoma.

The middle interscalene block: cadaver study and clinical assessment.

BACKGROUND AND OBJECTIVES: A variety of brachial plexus block techniques via the interscalene approach have been proposed. We describe here a new middle interscalene perivascular approach to the brachial plexus. To verify its effectiveness, we studied 719 patients scheduled for shoulder arthroscopy. Furthermore, to verify the accuracy of the proposed bony landmarks to use in the case of inability to palpate the subclavian artery pulse, we simulated the block on 10 cadavers. METHODS: The aim of our technique is to cannulate the neurovascular bundle by inserting a 35-mm needle lateral to the subclavian arterial pulse near the midpoint of the upper edge of the clavicle in a horizontal or slightly cephalad direction while pointing toward the seventh cervical vertebra. If the pulse of the subclavian artery is not palpable, we localize the direction of the needle with reference to 3 bony landmarks (the middle point of the clavicle, the spinous process of C7, and the sternoclavicular joint). By connecting these 3 landmarks, we obtain an angle whose apex lies at the midpoint of the clavicle and its bisecting line points to the plexus. The needle is introduced in the transverse plane of C7. RESULTS: The block was performed successfully in 692 of 719 cases (96.2%). Horner's syndrome occurred in 93.5% of the cases, arterial puncture with hematoma occurred in <1%, phrenic nerve block without respiratory impairment in 60%, with transient respiratory failure in <1%, and laryngeal nerve block in <1%. The incidence of severe complications or permanent injuries was zero (upper limit 95% confidence interval = 0.4% or 1:250 patients). The technique performed on cadavers showed that the previously mentioned bony landmarks were reliable reference points in reaching the brachial plexus. CONCLUSIONS: Our technique via a middle interscalene approach is easy to perform and provides a high success rate. Even in the absence of a subclavian artery pulse, the easily recognizable bony landmarks reliably guide us in the insertion of the needle. Furthermore, this technique appears to avoid complications that are theoretically possible in other supraclavicular perivascular approaches (pneumothorax) and paravertebral approaches (injection into the vertebral artery and subarachnoidal injection). However, further comparative studies will be required to assess the clinical relevance of the block.

Fatal interstitial pneumonitis associated with docetaxel administration in a patient with hormone-refractory prostate cancer.

Taxanes are widely used chemotherapeutic agents with the potential to induce pulmonary injury through a variety of mechanisms. Patients receiving these agents are at risk of acute or subacute pulmonary damage. The case is presented of a 72-year-old man with hormone-refractory prostate cancer and weekly administration of 30 mg/m2 docetaxel who developed subacute interstitial pneumonitis-related pulmonary fibrosis after seven doses and died despite mechanical ventilation and high-dose corticosteroid treatment. Even though only a few cases of this adverse event have been reported in the literature, severe docetaxel-induced pulmonary toxicity needs to be considered in the differential diagnosis when such patients present with respiratory symptoms.

The pathology of the spinal cord in progressive supranuclear palsy.

We describe the results of a study of the spinal cord of 5 patients with progressive supranuclear palsy (PSP). Examination of the 6th cervical, 7th thoracic, and 5th lumbar segments revealed variable degree of gliosis and density of neuropil threads (NTs), nerve cell loss, and tau-positive cytoplasmic staining of neurons, some of which was reminiscent of neurofibrillary tangles (NFT). Tau-positive neurons were seen at each spinal level and in the 3 zones in which each level was subdivided. Cells with the appearance of NFT predominated in the intermediate zone. Morphometric study revealed 47%, 52%, and 32% decrease in cell numbers in the motor area (lamina IX) at the 3 spinal levels, respectively, and 39% in the intermedio-lateral column. This is the first report describing severe neuronal loss throughout the whole spinal cord in patients with PSP and its results are in keeping with a previous study of the nucleus of Onufrowicz. The reasons why cell loss fails to produce clinical symptoms are analyzed and the clinico-pathological correlations between anatomical changes and dystonia are considered. On the basis of existing data, we conclude that previous suggestions implicating spinal interneurons in the pathogenesis of neck dystonia should not be supported.

p16INK4a is a useful marker for the diagnosis of adenocarcinoma of the cervix uteri and its precursors: an immunohistochemical study with immunocytochemical correlations.

p16 is a tumor suppressor gene that plays a central role in the regulation of the cell cycle. In squamous cervical cancers, overexpression of p16 is induced by HPV and associated with the carcinogenesis of cervical epithelia. The aim of this study was to determine whether immunostaining of p16 is useful in detecting adenocarcinomas of the cervix uteri in histologic and cytologic routine specimens. A total of 45 surgical specimens, including 18 cases of invasive carcinoma, 8 cases of adenocarcinoma in situ, 4 cases of endocervical glandular atypia (cervical glandular intraepithelial neoplasia), and 15 reactive lesions of the endocervical glands were immunostained using a specific anti-human p16 monoclonal antibody (clone E6H4, mtm laboratories AG, Heidelberg, Germany). Furthermore, immunocytochemical analysis was performed on 10 preoperative ThinPrep cytologic samples with abnormal glandular cells and compared with the human papillomavirus status as assessed with the Hybrid Capture II test. p16 was detected immunohistochemically in all 26 cases of adenocarcinoma of the cervix uteri, including 18 invasive and 8 in situ carcinomas. Only a focal expression was evidenced in the four specimens with endocervical glandular atypia, and no reaction was found in reactive lesions. Also, the immunocytochemical analysis on the 10 ThinPrep specimens evidenced a strong expression of p16 in neoplastic endocervical cells. In all cases this was associated with a high-risk HPV-positive typing. p16 is a useful marker for the detection of the adenocarcinoma of the cervix uteri and its precursors. The immunocytochemical detection on ThinPrep specimens may contribute to an early detection of endocervical lesions.

Human papillomavirus typing with hybrid capture II on archived liquid-based cytologic specimens: is HPV typing always reproducible?

Reproducibility of human papillomavirus (HPV) typing on archived ThinPrep (Cytyc, Boxborough, MA) specimens was evaluated repeating Hybrid Capture II (HCII) (Digene, Gaithersburg, MD) testing after 25 to 40 months (mean, 31.3 months; group 1), 6 to 11 months (mean, 8.4 months; group 2), and 0 to 5 months (mean, 3.5 months; group 3). Another ThinPrep slide was prepared to evaluate cellularity and reproducibility of the cytologic diagnosis. The mean residual relative light units (RLU) calculated for each group showed a strong decrease of RLU values at the second typing (group 1, 21.69%; group 2, 26.47%; and group 3, 32.25% of original values). No residual HPV DNA was shown in group 1 in 8 (13%) of 60 cases or in groups 2 and 3 in 2 (3%) of 60 cases each. These cases were associated mostly with poor cellularity and reproducibility of the initial cytologic diagnosis in the final cytologic examination. Intergroup statistical analysis of mean relative percentages for cases with satisfactory residual cellularity revealed a significant difference only between groups 1 and 3 (P < .05). Although mostly reproducible, HPV typing results by HCII on archived specimens are influenced by material consumption. In addition, results might be affected by some DNA degradation after long-term sample storage.

p16INK4a expression and progression risk of low-grade intraepithelial neoplasia of the cervix uteri.

The aim of the study was to evaluate the immunohistochemical expression of p16INK4a as a marker of progression risk in low-grade dysplastic lesions of the cervix uteri. p16INK4a immunohistochemistry was performed on 32 CIN1 with proven spontaneous regression of the lesion in the follow-up (group A), 31 (group B) with progression to CIN3 and 33 (group C) that were randomly chosen irrespective of the natural history of the lesion. p16INK4a staining pattern was scored as negative (less than 5% cells in the lower third of dysplastic epithelium stained), as focally positive (< or = 25%) and as diffuse positive (> 25%). A diffuse staining pattern was detected in 43.8% of CIN1 of group A, 74.2% of group B and 56.3% of group C. No p16INK4a staining was detected in 31.3% and 12.9% CIN1 lesions of groups A and B, respectively. Overall, 71.4% and 37.8% of p16INK4a-negative and diffusely positive CIN1 had regressed at follow-up, whereas 28.6% and 62.2% negative and diffusely positive CIN1 were progressed to CIN3, respectively (P < 0.05). All CIN3 lesions analyzed during follow-up of group B were diffusely stained for p16INK4a. Although p16INK4a may be expressed in low-grade squamous lesions that undergo spontaneous regression, in this study, CIN1 cases with diffuse p16INK4a staining had a significantly higher tendency to progress to a high-grade lesion than p16INK4a-negative cases. p16INK4a may have the potential to support the interpretation of low-grade dysplastic lesions of the cervix uteri.

Parasitism of the Zweeloo Woman: Dicrocoeliasis evidenced in a Roman period bog mummy.

We undertook the analysis of Zweeloo Woman, a bog mummy from the Netherlands, to assess her parasitic state. Evidence of infection came from two areas: (1) liver paraffin sections and (2) microfossils washed from an intestinal section. Although the liver had shrunken considerably, objects consistent with operculated trematode eggs were found. After evaluating the range of trematode species that produce eggs in liver tissue, we arrived at the diagnosis of Dicrocoelium dendriticum. Although only 0.1ml of sediment was recovered from an intestinal section, eggs of Ascaris lumbricoides and Trichuris trichiura were also identified. No eggs of D. dendriticum were revealed by the intestinal wash although they were observed in the liver. The lancet fluke, D. dendriticum, is a zoonosis that usually infects ruminants such as cattle. Eggs of D. dendriticum may be found in human coprolites if infected cow liver, for example, was eaten. This is false parasitism. Since eggs of D. dendriticum were found in the liver of Zweeloo Woman, we are assured this was a true infection. This find is especially significant because it is the oldest known, patent infection of D. dendriticum in humans.

Non-episodic angioedema associated with eosinophilia following Mycoplasma pneumoniae infection.

Episodic angioedema with eosinophilia is characterized by recurrent angioedema, peripheral eosinophilia, fever, weight gain, elevated serum immunoglobulin M (IgM), and a benign course lacking any internal organ involvement. A non-episodic variant has also been reported which is limited to a single attack and normally is less severe than the episodic type. We report a case of Mycoplasma pneumoniae infection with dermatological manifestation that was followed by non-episodic angioedema with eosinophilia including fever, weight gain, and elevated serum IgM. Even though the patient's clinical characteristics resemble episodic angioedema with eosinophilia as reported by Gleich, angioedema was non-episodic. This may be due to systemic corticosteroid treatment which was prescribed because of persistent skin manifestation following M. pneumoniae infection. The current report is the first observation suggesting that angioedema associated with eosinophilia may be triggered by atypical bacterial infection.

p16 ink4a and HPV L1 immunohistochemistry is helpful for estimating the behavior of low-grade dysplastic lesions of the cervix uteri.

As only a minority of low-grade dysplastic lesions of the cervix uteri will eventually progress to carcinoma, predicting the behavior of these lesions could be of high value in clinical practice. The aim of the study was to evaluate p16 ink4a and L1 as immunohistochemical markers of the biologic potentiality of low-grade dysplasia of the uterine cervix. The study included 38 conization specimens with coexisting cervical intraepithelial neoplasia grade 1 (CIN1) and 3 (CIN3) (group A) and 28 punch biopsies from women with CIN1 and proven spontaneous regression in the follow-up (group B). In group A, all CIN3 were p16 ink4a positive (p16+) and L1 negative (L1-). The CIN1 of this group were p16+L1- and p16+L1+ in 68.42% and 31.57%, respectively. No other expression pattern was found in this group. In group B, the p16+L1-, p16+L1+, p16-L1+, and p16-L1- patterns were found in 3.57%, 25%, 14.29%, and 57.14%, respectively. Overall, 96.29% p16+L1- CIN1 were found in group A, whereas all the p16-L1+ and p16-L1- CIN1 were found in group B. A significant difference between staining pattern distributions of group A and B was observed (P<0.0001). The results of the study show that p16 ink4a and L1 immunohistochemistry can be helpful for estimating the biologic potentiality of low-grade squamous cervical lesions. Particularly in cases in which the grade of the lesion is morphologically difficult to assess, the p16/L1 expression pattern could be useful for planning the clinical management of these women.

Abnormal cervicovaginal cytology with negative human papillomavirus testing.

BACKGROUND: Human papillomavirus (HPV) testing has become part of cervicovaginal diagnostics in many laboratories. False-negative HPV results, even if rare, are potentially relevant clinically, particularly when testing is used alone. In this study, the authors investigated the occurrence and causes of abnormal cervicovaginal cytologies with negative HPV testing. METHODS: The study was performed on 4130 liquid-based cytology (LBC) specimens from 2918 women who had abnormal cervical cytology or colposcopy or a history of abnormal cervical smear. HPV testing with Hybrid Capture II was repeated in LBC specimens with obviously atypical cytology (squamous intraepithelial lesion or abnormal squamous cells, cannot exclude high-grade lesion) and initial negative test results. The results were compared with the follow-up results and with the cytologic features of the specimens. RESULTS: Of 4130 LBC specimens, 94 (2.28%) showed obviously abnormal cells despite negative HPV results, and 50 of those 94 samples (53.19%) were HPV-positive on a repeated testing of the same specimen. Histologic follow-up showed high-grade intraepithelial lesion or carcinoma in 10 specimens that initially were negative for HPV. Eight of those samples were HPV-positive on testing repetition. Both specimens that had persistently negative results had poor-quality cytologic samples at revision. CONCLUSIONS: High-grade lesions may be identified in the follow-up of HPV-negative specimens with abnormal cytology. Testing repetition in patients with discordant cytology may prevent most potentially clinically relevant false-negative HPV results.