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The human HRAS1 belongs to an evolutionarily-conserved family of genes which enrolls among its members the yeast RAS2, a gene which regulates stress response and longevity in the Saccharomyces cerevisiae. In this paper we report that the frequency of the a3 allele of HRAS1 3'variable number tandem repeat (HRAS1 3'VNTR) decreases in centenarians in respect to young people, and we estimate that during aging a3 carriers have a relative mortality risk of 1.126 (95% CI=1.044-1.213). We propose that the germ-line variability at the HRAS1 locus impacts on the individual's capacity to reach the extreme limits of human life-span. Furthermore, we provide suggestive evidence that a3 HRAS1 3'VNTR allele and inherited variants of the mitochondrial genome (mtDNA haplogroups) do not affect independently human longevity, thus recalling the nucleus-mitochondrion interaction which regulates stress response and life-span in the yeast.

In this study, we introduce a centenarian-only approach to the assessment of gene-gene interaction that contributes to human longevity. This approach corresponds to the non-traditional case-only method in the genetic study of gene and disease associations. We first describe how the method can be implemented to screen for gene-gene interaction in human longevity. Then we apply the method to centenarian data collected from an Italian centenarian study in order to detect the interactions between the REN gene and the mitochondrial haplotypes. A significant interaction between REN gene allele 10 and the mitochondrial H haplotype, which may favour longevity, was found. Important features of the application in human longevity studies are highlighted and discussed. Since centenarians constitute a special population representing successful ageing, the centenarian-only approach will be an important tool in the search for major genes that contribute to human longevity.

Previous studies have shown that mitochondrial DNA (mtDNA) haplogroup J is significantly over-represented in healthy centenarians with respect to younger controls, thus suggesting that this haplogroup predisposes to successful aging and longevity. On the other hand, the same haplogroup is reported to have elevated frequency in some complex diseases. To verify if centenarians clustered in a particular lineage within J we have sequenced the D-loop region from 18 centenarians and 18 younger controls, previously characterized to be J. Then the entire mtDNA molecule was sequenced in a sub-sample of nine centenarians to find possible functional mutations associated with haplogroup J in successful aging. No clustering of the J haplogroup mtDNA from centenarians was observed. In addition, most of the mutations found are known as disease-associated mutations. The general picture that emerges from the study is that the J haplogroup of centenarians is surprisingly similar to that found in complex diseases, as well as in Leber Hereditary Optic Neuropathy. This finding implies that the same mutations could predispose to disease or longevity, probably according to individual-specific genetic backgrounds and stochastic events. This data reveals another paradox of centenarians and confirms the complexity of the longevity trait.

This paper reviews the recent literature on genes and longevity. The influence of genes on human life span has been confirmed in studies of life span correlation between related individuals based on family and twin data. Results from major twin studies indicate that approximately 25% of the variation in life span is genetically determined. Taking advantage of recent developments in molecular biology, researchers are now searching for candidate genes that might have an influence on life span. The data on unrelated individuals emerging from an ever-increasing number of centenarian studies makes this possible. This paper summarizes the rich literature dealing with the various aspects of the influence of genes on individual survival. Common phenomena affecting the development of disease and longevity are discussed. The major methodological difficulty one is confronted with when studying the epidemiology of longevity involves the complexity of the phenomenon, which arises from the polygenic nature of life span and historical mortality change. We discuss this issue and suggest new methodological approaches.

The role of interleukin 1 (IL-1) and interleukin 6 (IL-6) in the pathogenesis of Alzheimer's disease (AD) is reviewed within the framework of "inflamm-aging", i.e., the characteristic chronic pro-inflammatory status which develops in old age, and neuroinflammation, i.e., the peculiar inflammatory process which is present in the brain of AD patients. In particular, the data suggesting that several IL-1 and IL-6 gene polymorphisms can contribute to the risk of developing AD are reviewed. The possibility as well as the difficulty in identifying a pro-inflammatory phenotype, and its importance for the prevention, diagnosis and therapy of AD and other age-related pathologies are discussed.

Allele epsilon4 of the nuclear APOE gene is a leading genetic risk factor for sporadic Alzheimer's disease (AD). Moreover, an allele-specific effect of APOE isoforms on neuronal cell oxidative death is known. Because of the role of the mitochondrial genome (mtDNA) in oxidative phosphorylation and oxidative stress, an interaction between APOE polymorphism and mtDNA inherited variability in the genetic susceptibility to sporadic AD can be hypothesized. We have explored this hypothesis by analyzing mtDNA germline variants (mtDNA haplogroups) in a sample of AD patients (213 subjects) genotyped for APOE and classified as APOE epsilon4 carriers and non-carriers. We found that the frequency distribution of mtDNA haplogroups is different between epsilon4 carriers and non-carriers (P=0.018), thus showing non-random association between APOE and mtDNA polymorphisms. The same analysis, carried out in two samples of healthy subjects (179 age-matched and 210 individuals aged more than 100 years), showed independence between epsilon4 allele and mtDNA haplogroups. Therefore, the APOE/mtDNA interaction is restricted to AD and may affect susceptibility to the disease. In particular, some mtDNA haplogroups (K and U) seem to neutralize the harmful effect of the APOE epsilon4 allele, lowering the epsilon4 odds ratio from statistically significant to non-significant values.

We have analyzed a sample of 40 centenarians and 116 young controls from Sardinia, with a set of new Y chromosome binary markers, to evaluate if Y chromosome genes are involved in the high prevalence of males among centenarian Sardinians (1/2 vs. 1/4 in other populations studied). The results indicate that none of the seven lineages that account for >97% of the Y chromosome diversity in Sardinia provide an advantage with respect to the extreme longevity. However, our results, although based on the male-specific Y chromosome polymorphisms, give a clear profile of the pattern of genetic variability in Sardinia. Indeed they indicate that the Sardinian population had two main founder populations that have evolved in isolation for at least the last 5,000 years. These findings set the stage for future studies on longevity and other complex traits in Sardinia., (Copyright 2001 S. Karger AG, Basel)

New approaches are needed to explore the different ways in which genes affect the human life span. One needs to assess the genetic effects themselves, as well as gene-environment interactions and sex dependency. In this paper, we present a new model that combines both genotypic and demographic information in the estimation of the genetic influence on life spans. Based on Cox's proportional hazard assumption, the model measures the risks for each gene as well as for gene-environment and gene-sex interactions, while controlling for confounding factors. A two-step MLE is introduced to obtain a non-parametric form of the baseline hazard function. The model is applied to genotypic data from Italian centenarian studies to estimate relative risks of candidate genes, risks due to interactions and initial frequencies of different genes in the population. Results from models that either do or do not take into consideration individual heterogeneity are compared. It is shown that ignoring the existence of heterogeneity can lead to a systematic underestimation of genetic effects and effects due to interactions.

A genetic perspective of human history in Europe was derived from 22 binary markers of the nonrecombining Y chromosome (NRY). Ten lineages account for >95% of the 1007 European Y chromosomes studied. Geographic distribution and age estimates of alleles are compatible with two Paleolithic and one Neolithic migratory episode that have contributed to the modern European gene pool. A significant correlation between the NRY haplotype data and principal components based on 95 protein markers was observed, indicating the effectiveness of NRY binary polymorphisms in the characterization of human population composition and history.

In population studies of aging, the data on genetic markers are often collected for individuals from different age groups. The idea of such studies is to identify "longevity" or "frailty" genes by comparing the frequencies of genotypes in the oldest and in the younger groups of individuals. In this paper we discuss a new approach to the analysis of such data. This approach, based on the maximum likelihood method, combines data on genetic markers with survival information obtained from standard demographic life tables. This method allows us to evaluate survival characteristics for individuals carrying respective candidate genes. It can also be used in the estimation of the effects of allele-area and allele-allele interaction, either in the presence or absence of hidden heterogeneity. We apply this method to the analysis of Italian data on genetic markers for five autosomal loci and mitochondrial genomes. Then we discuss basic assumptions used in this analysis and directions of further research.

In this paper we extend the "network theory of aging," and we argue that a global reduction in the capacity to cope with a variety of stressors and a concomitant progressive increase in proinflammatory status are major characteristics of the aging process. This phenomenon, which we will refer to as "inflamm-aging," is provoked by a continuous antigenic load and stress. On the basis of evolutionary studies, we also argue that the immune and the stress responses are equivalent and that antigens are nothing other than particular types of stressors. We also propose to return macrophage to its rightful place as central actor not only in the inflammatory response and immunity, but also in the stress response. The rate of reaching the threshold of proinflammatory status over which diseases/disabilities ensue and the individual capacity to cope with and adapt to stressors are assumed to be complex traits with a genetic component. Finally, we argue that the persistence of inflammatory stimuli over time represents the biologic background (first hit) favoring the susceptibility to age-related diseases/disabilities. A second hit (absence of robust gene variants and/or presence of frail gene variants) is likely necessary to develop overt organ-specific age-related diseases having an inflammatory pathogenesis, such as atherosclerosis, Alzheimer's disease, osteoporosis, and diabetes. Following this perspective, several paradoxes of healthy centenarians (increase of plasma levels of inflammatory cytokines, acute phase proteins, and coagulation factors) are illustrated and explained. In conclusion, the beneficial effects of inflammation devoted to the neutralization of dangerous/harmful agents early in life and in adulthood become detrimental late in life in a period largely not foreseen by evolution, according to the antagonistic pleiotropy theory of aging.

Increasing data indicate that polymorphic variants of nuclear loci can affect rate and quality of aging in humans. However, the mitochondrial genome is another good candidate, because of the central role played by mitochondrial genes in oxidative phosphorylation (OXPHOS) and cell metabolism. A characteristic of the mitochondrial genome (mtDNA) is the high level of interindividual variability that ensues from high mutation rate and unilinear inheritance. Related groups of germline/inherited mtDNA polymorphisms (haplogroups) have been identified as continent-specific sets of stable/ancient/associated restriction fragment length polymorphisms in the mtDNA coding region, representing markers capable of exactly depicting the mtDNA pool of a specific population. The hypothesis can be put forward that mtDNA variants included in a haplogroup may have similar OXPHOS efficiency and therefore act as genetic factors predisposing to individual successful or unsuccessful aging. This idea can be explored by sampling groups of individuals of different ages from a well-defined population and comparing the pools of mtDNA haplogroups between samples. The results obtained by screening mtDNA haplogroups in about 800 Italians of different ages, including more than 200 centenarians, agree with the hypothesis that the inherited variability of the mitochondrial genome is associated with the chance of successful aging and longevity in humans.

Gender accounts for important differences in the incidence and prevalence of a variety of age-related diseases. Considering people of far advanced age, demographic data document a clear-cut prevalence of females compared to males, suggesting that sex-specific mortality rates follow different trajectories during aging. In the present investigation, we report data from a nationwide study on Italian centenarians (a total of 1162 subjects), and from two studies on centenarians living in two distinct zones of Italy, i.e., the island of Sardinia (a total of 222 subjects) and the Mantova province (Northern Italy) (a total of 43 subjects). The female/male ratio was about 2:1 in Sardinia, 4:1 in the whole of Italy, and about 7:1 in the Mantova province. Thus, a complex interaction of environmental, historical and genetic factors, differently characterizing the various parts of Italy, likely plays an important role in determining the gender-specific probability of achieving longevity. Gender differences in the health status of centenarians are also reported, and an innovative score method to classify long-lived people in different health categories, according to clinical and functional parameters, is proposed. Our data indicate that not only is this selected group of people, as a whole, highly heterogeneous, but also that a marked gender difference exists, since male centenarians are less heterogeneous and more healthy than female centenarians. Immunological factors regarding the age-related increase in pro-inflammatory status, and the frequency of HLA ancestral haplotypes also show gender differences that likely contribute to the different strategies that men and women seem to follow to achieve longevity. Concerning the different impact of genetic factors on the probability of reaching the extreme limits of the human life-span, emerging evidence (regarding mtDNA haplogroups, Thyrosine Hydroxilase, and IL-6 genes) suggests that female longevity is less dependent on genetics than male longevity, and that female centenarians likely exploited a healthier life-style and more favorable environmental conditions, owing to gender-specific cultural and anthropological characteristics of the Italian society in the last 100 years.

In population studies on aging, the data on genetic markers are often collected for individuals from different age groups. The purpose of such studies is to identify, by comparison of the frequencies of selected genotypes, "longevity" or "frailty" genes in the oldest and in younger groups of individuals. To address questions about more-complicated aspects of genetic influence on longevity, additional information must be used. In this article, we show that the use of demographic information, together with data on genetic markers, allows us to calculate hazard rates, relative risks, and survival functions for respective genes or genotypes. New methods of combining genetic and demographic information are discussed. These methods are tested on simulated data and then are applied to the analysis of data on genetic markers for two haplogroups of human mtDNA. The approaches suggested in this article provide a powerful tool for analyzing the influence of candidate genes on longevity and survival. We also show how factors such as changes in the initial frequencies of candidate genes in subsequent cohorts, or secular trends in cohort mortality, may influence the results of an analysis.

Mitochondrial DNA (mtDNA) is characterized by high variability, maternal inheritance, and absence of recombination. Studies of human populations have revealed ancestral associated polymorphisms whose combination defines groups of mtDNA types (haplogroups) that are currently used to reconstruct human evolution lineages. We used such inherited mtDNA markers to compare mtDNA population pools between a sample of individuals selected for successful aging and longevity (212 subjects older than 100 years and in good clinical condition) and a sample of 275 younger individuals (median age 38 years) carefully matched as to sex and geographic origin (northern and southern Italy). All nine haplogroups that are typical of Europeans were found in both samples, but male centenarians emerged in northern Italy as a particular sample: 1) mtDNA haplogroup frequency distribution was different between centenarians and younger individuals (P=0.017 by permutation tests); and 2) the frequency of the J haplogroup was notably higher in centenarians than in younger individuals (P=0.0052 by Fisher exact test). Since haplogroups are defined on the basis of inherited variants, these data show that mtDNA inherited variability could play a role in successful aging and longevity.

To identify possible genetic factors affecting human longevity we compared allele pools at two candidate loci for longevity between a sample of 143 centenarians (S) and a control sample of 158 individuals (C). The candidate loci were APOB and TPO, which code for apolipoprotein B and thyroid peroxidase, respectively. Both restriction fragment length (RFL) (XbaI2488 and EcoRI4154) and variable number of tandem repeat (VNTR) (3'APOB-VNTR) polymorphisms were analysed at the APOB locus; the TPO-VNTR polymorphism (intron 10) was analysed at the TPO locus. The main result of the investigation was that there is an association between the APOB locus and longevity that is revealed only when multiallelic polymorphisms are considered. In particular: (i) the frequency of 3'APOB-VNTR alleles with fewer than 35 repeats is significantly lower in cases than in controls; (ii) the linkage disequilibrium between the XbaI-RFLP and the EcoRI-RFLP is significantly different from 0 in cases but not in controls; (iii) the EcoRI-RFLP and XbaI-RFLP allele frequencies do not discriminate between cases and controls. The differences observed between case and control allele pools are specific to the APOB locus, since no significant difference was observed at the TPO locus.

Genotype and allele frequencies at seven Variable Number of Tandem Repeats (VNTR) loci currently used for forensic purposes have been estimated in a population sample from Calabria (south Italy). DNA target regions relevant to four microsatellites (THO.1; REN.4; D12S67; DYS19) and three minisatellites (D1S80; 3'APOB; TPO.10) were amplified by Polymerase Chain Reaction (PCR) and analysed by electrophoresis and ethidium bromide or silver staining. For all loci, the observed genotypes were found to be in agreement with those expected by the Hardy-Weinberg equilibrium. Data on allele frequencies were in line with those found in sample groups from northern or central Italy, tested for some of the above polymorphisms.

Linkage disequilibria in the apolipoprotein B (APOB) gene (EcoRI RFLP/3' APOB VNTR) and in the thyroid peroxidase (TPO) gene (AcyI RFLP/TPO VNTR) were investigated in a sample of 100 individuals from southern Italy. By recoding multiallelic data as diallelic data, each RFLP-VNTR system showed linkage disequilibrium significantly different from zero (EcoRI RFLP/3' APOB VNTR: p < 0.001; AcyI RFLP/TPO VNTR: p < 0.025), thus suggesting that the VNTR arrays are stable. Furthermore, the relationship between the 3' APOB (2p24-p23) and TPO (2pter-p24) VNTR multiallelic systems was also analyzed. The two VNTR polymorphisms were found to be in linkage equilibrium, thus indicating that they can be used together in forensic casework.

Genotype and allele frequencies of the DYS19, D12S67, and D1S80 highly polymorphic loci were determined in population samples from southern Italy (103 subjects) and Greece (84 subjects) using the amplified fragment length polymorphism (AFLP) technique (polymerase chain reaction followed by native PAGE and silver staining). Five, eleven, and eighteen alleles were found at the DYS19, D12S67, and D1S80 loci, respectively. PIC values ranged from 0.55 (DYS19 locus in Italians) to 0.79 (D12S67 locus in Italians). The distribution of D12S67 and D1S80 genotypes conformed to Hardy-Weinberg equilibrium, as confirmed by three statistics. Heterogeneity G tests, carried out on allele frequency distributions, showed a significant difference between the samples at the DYS19 locus, whereas no difference was found with regard to the other polymorphisms. Using data from the literature, we widened the comparison to other European groups analyzed for the same markers. All the polymorphisms were found to distinguish between populations of the same main ethnic group. In particular, D1S80 allele frequencies distinguished the Finns from other European groups (Spanish, German, Italian, and Greek samples). The reduced assay time, the high polymorphism level, and the ability to distinguish between populations indicate that these markers have potential value in population genetic studies.

Restriction fragment length polymorphisms (RFLPs) at codons 2488 (XbaI), 3611 (MspI), and 4154 (EcoRI) of the apolipoprotein B gene were investigated in sample groups from Athens (Greece) and Calabria (southern Italy) to verify whether the distribution of the APOB gene variants in Calabria, where Greek colonization occurred in the eighth century B.C., reflects that of the present Greek population. A sample from Apulia, a southern Italian region having a history different from that of Calabria, was also analyzed. Three specific DNA regions, each containing the polymorphic site, were amplified by polymerase chain reaction on 243 samples, and the restriction data for the three groups were compared. The allelic frequencies of the samples from Apulia and Greece showed variability patterns that agree with those found in Caucasians, whereas the Calabrian sample shows remarkable peculiarities, mainly for the EcoRI RFLP. Linkage disequilibrium analyses of pairs of markers showed strong D linkage values between X-M markers, whereas the D linkage values between M-R markers were too small to be reliably estimated. Last, for both Apulians and Greeks, X-R markers showed linkage disequilibrium, whereas for Calabrians they did not. Estimates of XMR haplotypic frequencies were computed; they were found to be appreciably different between Calabrian and Greek samples, whereas the frequencies in the Apulian sample were approximately midway between those in Calabrians and Greeks.

In order to characterize medulloblastomas and to get over the difficulties sometimes encountered in differential diagnosis, a double morphometric procedure has been applied to its nuclei. The first consisted of size measurements (maximum diameter, area and perimeter), the latter is represented by S.A.M. (Shape Analytical Morphometry) software-system specifically implemented to describe shape of biological structure by analytical parameters. Analytical and dimensional parameters submitted to Hotelling's multivariate discriminant analysis gave the best results when used together in convenient discriminant subsets, thereby allowing a good distinction between medulloblastoma in comparison with neuroblastoma, Ewing's tumor, lymphoblastic and lymphocytic lymphoma. These results underline the usefulness of morphometric characterization also for practical diagnostic purposes.

Recently, a method similar to ER.ICA has been proposed for the progesterone receptor (PgR) using two monoclonal antibodies, JZB39 and KD68, specific for human PgR and characterized by a molecular weight of 95 and 120 Kd, respectively. A series of 73 breast cancer patients was studied with regards to ER and PgR using both immunocytochemical (ICA) and biochemical (DCC) assays. Results showed no substantial differences between the two methods when considering common clinical-pathological parameters. Overall agreement between ICA and DCC methods was found: 79% for PgR and 78% for ER. A slight quantitative correlation was also observed between the "score values" of the ICA method and the Fmol content of ER and PgR using the Brave-Pearson test (r = 0.49 for PgR; r = 0.43 for ER). Specificity of PgR.ICA method was 77% for PgR and 72% for ER; sensitivity was 82% and 83%, respectively. The ICA method is a reliable technique to assess PgR presence as well as ER. Further studies are necessary to evaluate the prognostic role of nuclear PgR.

EcoRI restriction analysis at codon 4154 of the Apo B gene was performed in a sample of 90 subjects from southern Italy (sample S), using total blood cell DNA amplified by PCR. A group of 46 subjects from northern Italy (sample N) was also investigated for comparison. Southern Italians showed an incidence of the R2 allele (absence of the cutting site) twice as high as that found in northern Italians (48 v. 21%). By ASPCR the mutation which abolishes the restriction site was confirmed as being G----A at the first base of the 4154 codon of the Apo B gene (Glu----Lys) in both S and N samples. By studying the variability of cholesterolaemia among different EcoRI genotypes in the S sample, it was estimated that the average effect of the R2 allele is to lower serum cholesterol by 8.5 mg/dl.

The XbaI Restriction Fragment Length Polymorphism (RFLP) of the APOB gene at codon 2488 was investigated in a sample group from Southern Italy (165 subjects), taken from a population characterized by a low average level of cholesterolemia. The X2 allele (presence of XbaI cutting site), that in several groups was found to be associated with increased cholesterolemia, showed in Southern Italians a frequency of 39% which is significantly (P much less than 0.001) lower than that found in the majority of the Caucasoid groups so far tested (50%). However, an analysis of both cholesterol and APOB serum levels performed in a sample of 82 subjects, homogeneous for sex and age did not reveal any significant association between lipidemic parameters and APOB-XbaI genotypes.

Correlations of malignancy grade, immunologic phenotype and angiogenic capacity were studied in B-cell non-Hodgkin's lymphomas grafted onto chorioallantoic membranes of chicken embryos. The angiogenic response elicited by the tumors was significantly greater than the response to normal lymph nodes, but it did not correlate with either the malignancy grade or the immunologic phenotype of the tumors. The elevated angiogenic capacity of neoplastic tissues is confirmed. The results also suggest that the extensive vasoproliferative response required by a rapidly growing tumor is not only controlled by the neoplastic cell population but, probably, by the host-response as well.

Immunoisoelectrofocusing (IIEF) reveals a microheterogeneity of human serum IgA controlled by an autosomal polymorphic gene, termed S. The microheterogeneity disappears when sialic acid is removed from serum glycoproteins by neuraminidase treatment. It can be postulated, therefore, that S encodes a sialyltransferase which attaches sialic acid at the outer prosthetic chains of IgA.

16 cases of secondary ovarian tumors observed from 1977 to 1986, and in which the primary neoplasms involved other organs outside the genital tract or the breast, are studied. The observations concerned 13 metastases of the gastro-intestinal tract cancer (6 typical Krukenberg tumors, 3 "tubular" Krukenberg tumors, and 4 non-Krukenberg tumors), 1 urinary bladder cancer, 1 pancreatic cancer, and 1 tumor of unknown origin (probably an amelanotic melanoma). The histological diagnosis was very difficult in patients with no evidence of the primary malignancy, or in patients in whom the metastases simulate a concomitant primary ovarian tumor. The characteristic histological findings, histochemistry, and immuno histochemistry may suggest a correct diagnosis in most of situations.

Restriction fragment length polymorphism (RFLP) of human mitochondrial DNA was analysed in a sample of 87 subjects from Apulia (South Italy) by the restriction enzymes HpaI, BamHI, HaeII, MspI, AvaII, and HincII using total blood cell DNA probed by human mtDNA from placenta. Five BamHI morphs were observed, two of which are new (BamHI-4 and -5), as well as one new AvaII morph (AvaII-28). The association BamHI-4/AvaII-28 enables us to demonstrate for the first time two polymorphic BamHI sites present together in the same mtDNA molecule. In agreement with historical data the Apulian sample has been shown to be the most heterogeneous Italian population so far tested.

Surface ovarian epithelium and that of related inclusion cysts were comparatively studied in two groups of patients: one group of 50 non-pregnant women without myometrial, endometrial, or cervical hyperplasia and/or neoplasia; a second group of 50 women surgically treated for endometrial cancer and without evidence of ovarian pathology. Papillomatosis, hyperplasia and tubaric, squamoid, endometrioid and mucinous metaplasia were more frequently present in ovarian surface epithelium or in related inclusion cysts in patients with endometrial carcinoma. These findings may be correlated with a hormonal oncogenic stimulus involved in the genesis of endometrial cancer and suggests a new "hormonal" hypothesis in the pathogenesis of common epithelial tumors of the ovary.

This paper presents data on the identification of an allotypic polymorphism carried on a beta-lipoprotein in human serum. The antigen, whose inheritance is investigated in 78 families, is coded by a polymorphic locus (As) existing in 4 allelic states. The products of these alleles exhibit a serological linear subtyping pattern with respect to the alloantisera recognizing the allotypes. The allelic frequencies prove different in healthy (beta beta) and thalassemic (beta beta thal) sample populations, thus suggesting linkage dysequilibrium between the recessive alleles of the As and beta loci.

Radial Immuno Diffusion performed by including in agar serum from a polytransfused Cooley patient (beta-thalassemic homozygous) allowed to identify a human serum antigen marker of an autosomal polymorphic locus. the antigen and its coding gene were named AS and AS respectively. Genetic tests indicate that the AS1 dominant allele codes for the presence of AS in the serum while the AS0 recessive allele codes for its absence. The frequency of the AS0/AS0 homozygous genotype results to be greater in Cooley than in healthy people thus suggesting that the AS0 and beta-thal alleles may be in linkage disequilibrium. The observation that the frequency of the AS0 beta-thal haplotype is greater than the product of the frequencies of the alleles which form it, confirms such a linkage.

This paper presents data on a human allotypic polymorphism carried on a low density lipoprotein (As-LDL). The As polymorphism was investigated by a rabbit antiserum which was firstly used to analyze the serum level and the isoelectric point of the As lipoprotein in sera of different phenotype. This antiserum was also used to purify As-LDL by immunochromatography. Afterwards the peptidic maps of the APO proteins from different As-LDL were compared. While no correlation did result between the serum level and the serological phenotype of the As lipoprotein, the peptidic distribution of the APO parts from different As-LDL showed differences correlated to the allotypic phenotype.

The paper describes a cattle serum antigen (LdlA1) located on a low-density lipoprotein and detected by single radial diffusion. The specificity is inherited in a simple Mendelian manner and the gene controlling its synthesis is inherited independently from the one controlling the synthesis of the alpha 2 macroglobulin McA1 antigen.

In the attempt to discriminate between centroblasts and immunoblasts, an analytical morphometrical procedure was adopted, considering the nuclei of a randomly selected centroblastic and immunoblastic population from non-Hodgkin's lymphomas and the nucleoli of a more restricted number of the same neoplastic population. All the size-independent shape-descriptor parameters extracted for each step of the mathematical analyses used were submitted to Hotelling's multivariate discriminant analysis that enabled us to achieve a good distinction (3% error) between centroblasts and immunoblasts when nuclear and nucleolar parameters were used together.

Breast cancer nodule shape has been investigated by two analytical procedures: upper degree polynomials and harmonic Fourier analysis. An automized method (TV camera interfaced with a computer) has been employed to find the coefficients of kth order equation giving the best fit. Then the difference between real node contour and function curve has been calculated and submitted to harmonic analysis. This gives the sine/cosine coefficients of the two series and the amplitude of the harmonics. The results point out that no difference exists between metastatic and non metastatic tumors and suggest that parametrized descriptors calculated may represent a peculiar behaviour in the growth of the tumor.

The aim of this study was to investigate the biological significance of tumor shape in breast cancer by considering the shape not as a casual event but as an expression of the behavior and natural history of the tumor. The shape was studied by an analytical morphometry procedure and was related to axillary metastases, which up to now are the most meaningful prognostic factors in this disease. Fifty cases of infiltrating breast carcinoma (25 N+ and 25 N-) were investigated. The shape, studied on subgross sections of the tumor, was considered as the result of two components: the subtle contour irregularities and the main distortions of the figure. The procedures used allowed us to distinguish and to parametrize these two components in order to submit them to univariate analysis (Student's t test), a principal components analysis and, finally, a multivariate discriminant analysis (Hotelling test). The utilized analytical procedures by work-station S.A.M. (S.A.M. work station is a product of "Metamorphosis") consisted of three steps: 1) Extraction of tumor function curve obtained by Kth order polynominals which gives a smoothing effect to the original curve; 2) Evaluation of contour irregularities by Fourier harmonic analysis; 3) Evaluation of shape asymmetry by SAE (shape asymmetry evaluator). We considered also the roundness factors of the original and function curves and finally the maximum tumor diameter. Three parameters relating to contour irregularities (sum and mean value of Fourier harmonic amplitude and percentage of superimposed points) and parameters relating to main distortions of the figure (mean value of SAE) were highly significant (p less than 0.001). The roundness factor of the original curve was more significant (p less than 0.001) than that of the function curve (p less than 0.01) whereas maximum tumor diameter was not significant when tested by Student's t test. Multivariate discriminant analysis allowed 20% of error to be reached by using 3 parameters relating to the shape analysis and the two roundness factors. By using 8 parameters, including maximum tumor diameter, the percentage error was 16%. The results obtained, while they stress the usefulness of the employed procedure, reveal that shape of the tumor, together with its dimensions, is an important expression of the biological behavior relating to metastatic spread.

An adult with clinically and neuroradiologically occult diastematomyelia is described. The neuropathological study revealed an unusual duplication of the spinal cord not associated with osteocartilagineous anomalies. This anomaly probably occurs early in fetal life as an isolated lesion of the neural tube. The embryological mechanism is discussed.

G1m(a) and G1m(x) allotype distribution is analyzed in a sample population from Calabria (South Italy). The paper presents data showing an unusually high frequency of the G1m(x) marker. Furthermore, results concerning a rare Gm(x) phenotype are also shown. The findings suggest the important contribution yielded to the Calabrian genetic pool by the Albanian population that immigrated to this region in the 15th century.

mtDNA polymorphisms were studied in a sample of 90 individuals of the Sicilian population using six restriction enzymes: HpaI, BamHI, HaeII, MspI, AvaII and HincII. (1) Three new patterns, for MspI, AvaII and HincII, have been detected. (2) At least two different mutations were found to account for both the AvaII morph 3 and the AvaII morph 9 as in many other Caucasian groups so far examined. (3) Seventeen types were found; of these six are new. The frequency (54.5%) of type 1-2 (2.1.1.1.1.2) is lower than in the rest of Italy whereas those of type 6-2 (2.1.2.1.1.2) (10.0%) and type 18-2 (2.3.1.4.9*.2) (12.2%) lie at the upper level of the Italian range. The 18-derivative, type 57-2 (2.3.1.4.13*.2), which is consistently found in all Italian samples, is present also among Sicilians with an incidence of 2.2%. (4) Of particular interest is that the HpaI-3/AvaII-3 complex, which is unique to groups of African ancestry, was found in Sicily at a frequency of 4.4%. For the first time an estimate of the amount of gene flow from Blacks to the Sicilian gene pool could be obtained.

Humoral proteins from Amphioxus lanceolatus showed a considerable agglutinating activity for human red cells. The haemagglutinating factor eluted in the macroglobulin peak on Sephadex G200 and resulted ConA-1 at affinity chromatography on Sepharose ConA. Immunoelectrophoresis, carried out using a specific antiserum produced in rabbit, revealed alpha mobility for the agglutinating factor, thus suggesting that it was an alpha macroglobulin.

The G1m(1) and G1m(2) allotype distribution was analyzed in a population sample from 11 Albanian towns of Calabria. The unusually high frequency of the G1m(1) marker already observed in Calabria as well as the presence of the Gm(2) phenotype were shown. The Calabrian and Albanian populations were similar, but significantly different from other Italian populations.