Expression of decorin throughout the murine hair follicle cycle: hair cycle dependence and anagen phase prolongation

Decorin is a prototypical member of the small leucine‐rich proteoglycan (SLRP) family, which is involved in numerous biological processes. The role of decorin, as a representative SLRP, in hair follicle morphogenesis has not been elucidated. We present our initial findings on decorin expression patterns during induced murine hair follicle (HF) cycles. It was found that decorin expression is exclusively restricted to the epidermis, outer root sheath and sebaceous glands during the anagen phase, which correlates with the upregulation of decorin mRNA and protein expression in depilated murine dorsal skin. Furthermore, we used a functional approach to investigate the effects of recombinant human decorin (rhDecorin) via cutaneous injection into HFs at various murine hair cycle stages. The local injection of rhDecorin (100 μg/ml) into the hypodermis of depilated C57BL/6 mice at anagen delayed catagen progression. In contrast, rhDecorin injection during the telogen phase caused the premature onset of anagen, as demonstrated by the assessment of the following parameters: (i) hair shaft length, (ii) follicular bulbar diameter, (iii) hair follicle cycling score and (iv) follicular phase percentage. Taken together, our results suggest that decorin may modulate follicular cycling and morphogenesis. In addition, this study also provides insight into the molecular control mechanisms governing hair follicular epithelial–mesenchymal interactions.     

Hair follicle expression of 1,25-dihydroxyvitamin D3 receptors during the murine hair cycle

Because the hair follicle is a highly hormone-sensitive miniorgan, the role of hormones produced locally in the skin in the control of hair growth deserves systematic analysis. It has been shown previously that the potent steroid hormone 1,25-dihydroxyvitamin D₃ (1,25-D₃) modulates growth and differentiation of keratinocytes via binding to a high-affinity nuclear vitamin D receptor (VDR). In this study, we have examined the in situ expression of VDR during the murine hair cycle. VDR expression was detected immunohistochemically. To obtain defined stages of the murine hair cycle, hair growth was induced by depilation in C57 BL-6 mice. In addition to the recognized VDR expression of outer root sheath keratinocytes, we detected VDR immunoreactive cells in the dermal papilla, the mesenchymal key structure of the hair follicle. Furthermore, VDR immunoreactivity in the nuclei of outer root sheath keratinocytes and in dermal papilla cells was stronger during anagen IV-VI and catagen than during telogen and anagen I-III. This suggests hair cycle-associated changes in the expression of VDR, and points to a potential role for 1,25-D₃ in hair follicle biology. Selected follicular cell populations may display hair cycle-dependent sensitivity to 1,25-D₃ stimulation.     

E- and P-cadherin expression during murine hair follicle morphogenesis and cycling

The role of adhesion molecules in the control of hair follicle (HF) morphogenesis, regression and cycling is still rather enigmatic. Since the adhesion molecules E- and P-cadherin (Ecad and Pcad) are functionally important, e.g. during embryonic pattern formation, we have studied their expression patterns during neonatal HF morphogenesis and cycling in C57/BL6 mice by immunohistology and semi-quantitative RT-PCR. The expression of both cadherins was strikingly hair cycle-dependent and restricted to distinct anatomical HF compartments. During HF morphogenesis, hair bud keratinocytes displayed strong Ecad and Pcad immunoreactivity (IR). While neonatal epidermis showed Ecad IR in all epidermal layers, Pcad IR was restricted to the basal layer. During later stages of HF morphogenesis and during anagen IV-VI of the adolescent murine hair cycle, the outer root sheath showed strong E- and Pcad IR. Instead, the outermost portion of the hair matrix and the inner root sheath displayed isolated Ecad IR, while the innermost portion of the hair matrix exhibited isolated Pcad IR. During telogen, all epidermal and follicular keratinocytes showed strong Ecad IR. This is in contrast to Pcad, whose IR was stringently restricted to matrix and secondary hair germ keratinocytes which are in closest proximity to the dermal papilla. These findings suggest that isolated or combined E- and/or Pcad expression is involved in follicular pattern formation by segregating HF keratinocytes into functionally distinct subpopulations; most notably, isolated Pcad expression may segregate those hair matrix keratinocytes into one functional epithelial tissue unit, which is particularly susceptible to growth control by dermal papilla-derived morphogens. The next challenge is to define which secreted agents implicated in hair growth control modulate these follicular cadherin expression patterns, and to define how these basic parameters of HF topobiology are altered during common hair growth disorders.     

真皮干细胞在毛囊周期中的作用

毛囊由表皮(上皮)及真皮(间充质)组成,它们之间的相互作用在毛囊的形态发生及生长中发挥重要作用,二者之间相互作用是毛囊成功重建的关键因素.在毛发形成过程中,真皮细胞是诱导者,上皮细胞是应答者.真皮鞘和毛乳头内存在毛囊真皮干细胞,属于成体干细胞,具有慢周期、未分化、自我更新和体外增殖能力强的特点.真皮鞘中的真皮干细胞较长寿,可以历经几个毛囊周期,重建真皮鞘.在毛囊周期的生长期,真皮鞘中的干细胞产生新细胞提供给毛乳头;在退行期,真皮干细胞子代移出毛乳头或死亡.毛囊真皮细胞对于损伤和疾病之后的毛囊重建及修复具有重要意义.     

Control of human hair growth by neurotrophins: brain-derived neurotrophic factor inhibits hair shaft elongation, induces catagen, and stimulates follicular transforming growth factor beta2 expression

Neurotrophins are important modulators of epithelial-mesenchymal interactions. Previously, we had shown that brain-derived neurotrophic factor (BDNF) and its high-affinity receptor tyrosine kinase B (TrkB) are prominently involved in the control of murine hair follicle cycling. We now show that BDNF and TrkB are also expressed in the human hair follicle in a manner that is both hair cycle dependent and suggestive of epithelial-mesenchymal cross-talk between BDNF-secreting dermal papilla fibroblasts of anagen hair follicles and subpopulations of TrkB+ hair follicle keratinocytes. As functional evidence for an involvement of BDNF/TrkB in human hair growth control, we show in organ-cultured human anagen hair follicles that 50 ng per mL BDNF significantly inhibit hair shaft elongation, induce premature catagen development, and inhibit keratinocyte proliferation. Quantitative real-time rtPCR analysis demonstrates upregulation of the potent catagen inducer, transforming growth factor beta2 (TGFbeta2) by BDNF, whereas catagen induction by BDNF was partially reversible through co-administration of TGFbeta-neutralizing antibody. This suggests that TrkB-mediated signaling promotes the switch between anagen and catagen at least in part via upregulation of TGFbeta2. Thus, human scalp hair follicles are both a source and target of bioregulation by BDNF, which invites to target TrkB-mediated signaling for therapeutic hair growth modulation.     

Hair cycle-specific immunolocalization of retinoic acid synthesizing enzymes Aldh1a2 and Aldh1a3 indicate complex regulation

Retinoic acid has long been known to alter skin and hair growth but an exact mechanism is unclear. This study was performed to examine the sites of endogenous retinoic acid synthesis in the cycling hair follicle to better understand the role retinoic acid plays in this process. Retinal dehydrogenases (Aldh1a1, 2, and 3, formerly Raldh 1, 2, and 3) are the enzymes responsible for the last step in retinoic acid synthesis. Immunohistochemistry was performed on adult C57BL/6J mouse skin sections with antibodies against Aldh1a2 and Aldh1a3. Aldh1a2 expression was seen primarily in the outer root sheath and basal/spinous layer during all stages of the hair cycle, and in the bulge during anagen and early catagen, whereas Aldh1a3 expression was primarily in the dermal papilla, pre-cortex, and hair shaft during mid-late anagen. The expression patterns of these two similar retinoic acid synthesizing enzymes at specific follicular sites suggest that they mediate and are regulated by different epithelial proliferation and differentiation signaling pathways.     

Transforming growth factor-beta receptor II is preferentially expressed in the companion layer of the human anagen hair follicle

BACKGROUND: Transforming growth factor (TGF)-beta is a multifunctional growth factor with multiple roles in skin including hair follicle development and cycling, where it regulates cell proliferation, differentiation and apoptosis, as well as in wound healing. While TGF-beta receptor I (TGF-beta RI) and receptor II (TGF-beta RII) expression helps define early human hair follicle morphogenesis, expression in the adult human hair follicle remains to be established. OBJECTIVES: To assess TGF-beta receptor expression in human scalp anagen hair follicles. METHODS: Immunohistochemical and double immunofluorescence analysis of TGF-beta RI and RII was conducted on frozen sections of haired human scalp obtained from 10 healthy individuals. RESULTS: TGF-beta RI expression was detected in the outer root sheath of anagen hair follicles while TGF-beta RII was expressed almost exclusively in the companion layer of inner root sheath and less so in premedulla keratinocytes. Both receptors were colocalized in the companion layer of the proximal and mid follicle. CONCLUSIONS: The well-described role of TGF-beta in keratinocyte apoptosis during catagen is likely to involve anagen-specific hair follicle components including the companion layer, as this layer provides the slippage plane supporting the inner root sheath and hair shaft as they ascend to the skin surface. Results of this study suggest that the colocalization of TGF-beta RI/RII complexes at the companion layer would facilitate TGF-beta signalling at this site to regulate apoptosis of the companion layer keratinocytes, facilitating shrinkage/contraction of this cell layer during hair follicle regression/catagen.     

Neuron-specific PGP9.5 expression in rat hair follicle development and cycle

Protein gene product PGP9.5 is a neuron-specific ubiquitin C-terminal hydrolase. We found that it also has immunoreactivity in the hair follicle of the Wistar rat dorsal skin and its expression patterns change with the development and cycle. During the morphogenesis, the PGP9.5 was expressed in the hair germ and hair peg elongated from the epidermis, and became restricted in the outer root sheath as the development progressed. In catagen, however, the PGP9.5 was detected in the tailing epithelial strand of the regressing proximal follicle epithelium, and in the keratinocytes directly contacted with the club hair, but rarely in the outer root sheath. With the beginning of the anagen of the second hair follicle, the PGP9.5 was again expressed in the second hair germ, and in the keratinocytes surrounding the remaining club hair and of distal follicle of the first hair. These findings showed that PGP9.5 is not specific to the neuron but is also involved in the hair follicle, and should provide new insight into the development and regression of the hair follicle.     

Plasticity and cytokinetic dynamics of the hair follicle mesenchyme: implications for hair growth control

The continuously remodeled hair follicle is a uniquely exploitable epithelial-mesenchymal interaction system. In contrast to the cyclical fate of the hair follicle epithelium, the dynamics of the supposedly stable hair follicle mesenchyme remains enigmatic. Here we address this issue using the C57BL/6 hair research model. During hair growth, increase in total follicular papilla size was associated with doubling of papilla cell numbers, much of which occurred before intra-follicular papilla cell proliferation, and subsequent to mitosis in the proximal connective tissue sheath. This indicates that some papilla cells originate in, and migrate from, the proliferating pool of connective tissue sheath fibroblasts. Follicular papilla cell number and total papilla size were maximal by anagen VI, but intriguingly, decreased by 25% during this period of sustained hair production. This cell loss, which continued during catagen, was not associated with intra-follicular papilla apoptosis, strongly indicating that fibroblasts migrate out of the late anagen/early catagen papilla and re-enter the proximal connective tissue sheath. Low-level apoptosis occurred only here, along with the "detachment" of cells from the regressing connective tissue sheath. Thus, the hair follicle mesenchyme exhibits significant hair cycle-associated plasticity. Modulation of these cell interchanges is likely to be important during clinically important hair follicle transformations, e.g. vellus-to-terminal and terminal-to-vellus during androgenetic alopecia.     

Immunohistochemical analysis of tissue remodelling during the anagen—catagen transition of the human hair follicle

The transition from the growth phase (anagen) to the involution phase (catagen) involves profound morphological changes in the human hair follicle. Club hair and epithelial column formation, for example, are key features of the catagen phase, which result in the disruption of physical interaction between the bulb and the dermal papilla. However, the dynamics and tissue remodelling that occur during this involution process remain largely unknown. Using monoclonal antibodies directed against K14 keratin, trichohyalin, transglutaminase I, desmoglein and Ki67 antigen, we followed the movements of each of the main hair follicle compartments during the onset of catagen. Our results indicate that the inner root sheath is an early target in this process, suggesting a key role for this compartment in the maintenance of hair follicle homeostasis.     

Molecular control of epithelial-mesenchymal interactions during hair follicle cycling

Epithelial-mesenchymal interactions play pivotal roles in the morphogenesis of many organs and various types of appendages. During hair follicle development, extensive interactions between two embryologically different hair follicle compartments (epidermal keratinocytes and dermal papilla fibroblasts) lead to the formation of the hair shaft-producing mini-organ that shows cyclic activity during postnatal life with periods of active growth, involution and resting. During the hair cycle, the epithelium and the mesenchyme are regulated by a distinct set of molecular signals that are unique for every distinct phase of the hair cycle. In telogen hair follicles, epithelial-mesenchymal interactions are characterized by a predominance of inhibitory signals that retain the hair follicle in a quiescent state. During anagen, a large variety of growth stimulatory pathways are activated in the epithelium and in the mesenchyme, the coordination of which are essential for proper hair fiber formation. During catagen, the termination of anagen-specific signaling interactions between the epithelium and the mesenchyme leads to apoptosis in the hair follicle epithelium, while activation of selected signaling pathways promotes the transition of the dermal papilla into a quiescent state. The signaling exchange between the follicular epithelium and the mesenchyme is modulated by proteoglycans, such as versican, which may significantly enhance or reduce the biological activities of secreted growth stimulators. However, additional research will be required to bridge the gap between our current understanding of mechanisms underlying epithelial-mesenchymal interactions in hair follicles and the potential clinical application of growth modulators involved in those interactions. Further progress in this area of research will hopefully lead to the development of new drugs for the treatment of hair growth disorders.     

The role of BMP signalling in the control of ID3 expression in the hair follicle

Both the production of the hair shaft in anagen and the initiation of a new hair cycle at telogen are the result of reciprocal interactions between the dermal papilla and the overlying epithelial cells. Secreted factors, such as those of the bone morphogenetic protein (BMP) family, play a crucial role in moderating these interactions. Analysis of hair follicles in different stages of the hair cycle showed that BMP signalling was only active during anagen and again during telogen. During catagen, no BMP signalling occurred in the dermal papilla. ID3, a gene expressed in the dermal papilla of both vibrissa and pelage follicles, is a BMP target, and as such, we found that ID3 was expressed from the earliest stages of morphogenesis. During the hair cycle, ID3 was only expressed in the dermal papilla at middle anagen and telogen. To test the significance of ID3 expression in the dermal papilla, we cultured dermal papilla cells and found that ID3 expression fell significantly after a single passage. ID3 expression was returned to in vivo levels in low- and high-passage cells by culturing to high confluence or by the addition of BMP4. These studies reinforce the requirement for active BMP signalling and cell-cell contacts in the dermal papilla during specific stages in the hair cycle.     

Dickkopf 1 promotes regression of hair follicles

Recently, we suggested that Dickkopf 1 (DKK-1) is a pathogenic mediator involved in male pattern baldness. As premature catagen onset is a key characteristic of male pattern baldness, in this study, we evaluated whether DKK-1 has a role as a catagen inducer in hair cycling. Herein, we report that recombinant human DKK-1 (rhDKK-1) injection into the hypodermis of mice during anagen caused premature onset of catagen, whereas neutralizing DKK-1 antibody delayed anagen-to-catagen transition in mice. Moreover, treatment with rhDKK-1 led to a decrease in final hair follicle length, whereas DKK-1 antibody led to an increase compared with control animals. In addition, DKK-1 and DKK-1 messenger RNA expression is most upregulated in follicular keratinocytes of late anagen in depilation-induced hair cycle progression. Moreover, we observed that rhDKK-1 blocks canonical Wnt-mediated activation of β-catenin signaling and induces the proapoptotic protein Bax, resulting in apoptosis in outer root sheath keratinocytes. Taken together, our data strongly suggest that DKK-1 is involved in anagen-to-catagen transition in the hair cycle by regulating the activity of follicular keratinocytes.     

Hair follicle pigmentation

Hair shaft melanin components (eu- or/and pheomelanin) are a long-lived record of precise interactions in the hair follicle pigmentary unit, e.g., between follicular melanocytes, keratinocytes, and dermal papilla fibroblasts. Follicular melanogenesis (FM) involves sequentially the melanogenic activity of follicular melanocytes, the transfer of melanin granules into cortical and medulla keratinocytes, and the formation of pigmented hair shafts. This activity is in turn regulated by an array of enzymes, structural and regulatory proteins, transporters, and receptors and their ligands, acting on the developmental stages, cellular, and hair follicle levels. FM is stringently coupled to the anagen stage of the hair cycle, being switched-off in catagen to remain absent through telogen. At the organ level FM is precisely coupled to the life cycle of melanocytes with changes in their compartmental distribution and accelerated melanoblast/melanocyte differentiation with enhanced secretory activity. The melanocyte compartments in the upper hair follicle also provides a reservoir for the repigmentation of epidermis and, for the cyclic formation of new anagen hair bulbs. Melanin synthesis and pigment transfer to bulb keratinocytes are dependent on the availability of melanin precursors, and regulation by signal transduction pathways intrinsic to skin and hair follicle, which are both receptor dependent and independent, act through auto-, para- or intracrine mechanisms and can be modified by hormonal signals. The important regulators are MC1 receptor its and adrenocorticotropic hormone, melanocyte stimulating hormone, agouti protein ligands (in rodents), c-Kit, and the endothelin receptors with their ligands. Melanin itself has a wide range of bioactivities that extend far beyond its determination of hair color.     

The fate of hair follicle melanocytes during the hair growth cycle

The fate of the follicular pigmentary unit during the hair growth cycle has long been one of the great enigmas of both hair follicle and pigment cell biology. Although melanocytes are distributed in several different compartments of the anagen hair follicle, melanogenically active cells are located only in the hair bulb, where they are directly involved in hair shaft pigmentation. These pigment cells are readily detectable only when they become melanogenically active during anagen III of the hair growth cycle. Thus, their status during hair follicle regression (catagen), when melanogenesis is switched off, until they re-appear again as pigment-producing cells in the anagen III hair follicle, has remained poorly defined. Historically, it has been proposed that hair bulb melanocytes adopt a self-perpetuating, catagen-resistant strategy of de-differentiation during hair follicle regression and re-differentiation upon entry into a new anagen phase; however, this explanation remains problematic in the absence of evidence for de-differentiation/re-differentiation plasticity in most nonmalignant cell systems.     

Expression of cyclooxygenase isozymes during morphogenesis and cycling of pelage hair follicles in mouse skin: precocious onset of the first catagen phase and alopecia upon cyclooxygenase-2 overexpression

Cyclooxygenase (COX)-1 and -2 catalyze the key reaction in prostaglandin biosynthesis. Whereas COX-1 is found in most tissues, COX-2, with a few exceptions, is not expressed in normal tissues but becomes transiently induced in the course of inflammatory reactions. In many neoplastic epithelia, COX-2 is constitutively overexpressed. Here we show that COX isozymes are spatiotemporally expressed during morphogenesis of dorsal skin epithelium of NMRI mice. COX-1 and COX-2 mRNA and protein were detected in embryonic and postnatal epidermal tissue by RT-PCR, northern blot, and immunoblot analysis indicating that both isoforms may contribute to prostaglandin production. Being barely detectable in interfollicular epidermis and resting hair follicles of adult mice, COX-2 protein appeared in embryonic skin first in epidermal precursor cells and later on in the basal cells and the peridermal layer of the stratified epidermis. In the course of pelage hair follicle morphogenesis, COX-2 remained expressed in the basal interfollicular compartment and, in addition, became apparent in elongated hair germs and hair pegs and later on in the outer root sheath cells of the distal and proximal hair follicles as well as in basal sebaceous gland cells. During the subsequent synchronous phases of hair cycling, COX-2 expression declined in catagen, was barely detectable in telogen, and was reinduced in the basal outer root sheath and basal sebaceous gland cells of anagen hair follicles. COX-1 immunosignals were detected predominantly in the interfollicular spinous and granular layers of the developing, neonatal, and adult epidermis but not in follicular epithelial cells of developing or cycling hair follicles. Dendritic cells in the interfollicular epidermis and distal hair follicles were also COX-1-positive. Transgenic overexpression of COX-2 under the control of a keratin 5 promoter in basal cells of the interfollicular and follicular epidermis induced a precocious entry into the first catagen stage of postnatal hair follicle cycling and a subsequent disturbance of hair follicle phasing. Furthermore, transgenic mice developed an alopecia. Inhibition of transgenic COX-2 activity by feeding the specific COX-2 inhibitor valdecoxib suppressed the development of alopecia, indicating that COX-2-mediated prostaglandin synthesis is involved in hair follicle biology.     

Interleukin-4 induces apoptosis in cultured human follicular keratinocytes,but not in dermal papilla cells

The importance of apoptosis in hair follicle cycling is still not clearly understood, however, its regulation in follicular keratinocytes (FK) during bulb regression (catagen) may be essential for hair regrowth. So far, the control of FK apoptosis remains unknown. In this study, the anti-inflammatory cytokine IL-4 was found to induce apoptosis dose and time dependently in cultured human FK, in contrast to other agents known to inhibit hair growth such as IL-1alpha, IL-1beta, TNFalpha and TGFbeta, as shown by DNA fragmentation. On the other hand, cytokines reported to be involved in hair follicle cycling including IL-4 were not able to induce apoptosis in dermal papilla cells (DPC), in contrast to staurosporine. This PKC inhibitor revealed dose-dependent apoptotic signals not only for DPC but also for FK in vitro. In further experiments the expression of apoptosis regulating proteins, possibly involved in catagen formation, was analyzed in FK and DPC. However, no striking difference in RNA expression was seen in either cell population under culture conditions and after incubation with IL-4. We conclude, therefore, that IL-4 mediated apoptosis may participate in regulating catagen formation in the hair follicle, acting selectively on cultured FK and being independent of bcl-2 and bax expression.     

Human scalp skin and hair follicles express neurotrophin-3 and its high-affinity receptor tyrosine kinase C, and show hair cycle-dependent alterations in expression

BACKGROUND: Neurotrophin (NT)-3 and its high-affinity receptor tyrosine kinase C (Trk C) are essential for nervous system development. These members of the NT family are also involved in murine hair morphogenesis and cycling. However, their role in human hair follicle (HF) biology remains to be elucidated. OBJECTIVES: To explore the role of NTs in human skin and HF biology. METHODS: The immunoreactivity (IR) of NT-3 and Trk C was studied in human scalp skin and HFs by immunofluorescent and light microscopic immunohistology. Skin biopsies were obtained from normal human scalp containing mainly anagen VI HFs from women (age 53-57 years) undergoing elective plastic surgery. RESULTS: Both NT-3 and Trk C showed prominent, yet distinct, IR patterns in human scalp anagen HFs (anagen VI), whereas they were weakly expressed in catagen and increased again in telogen HFs. Within HF compartments, NT-3 IR was prominent in the outer root sheath, inner root sheath, dermal papilla and connective tissue sheath. Trk C IR was prominent in all HF epithelial and mesenchymal compartments. Outside the HF, both NT-3 and Trk C showed prominent IR in the epidermis, sebaceous glands and sweat glands. CONCLUSIONS: These observations provide the first indication that NT-3 and Trk C are expressed in human scalp skin and HFs, and suggest that Trk C-mediated signalling is involved not only in murine but also in human HF biology. They may be useful in determining therapeutic strategies for the treatment of hair cycle and skin-related disorders.     

Hair cycle-dependent expression of heat shock proteins in hair follicle epithelium

Background Heat shock proteins (HSPs) have a physiologic function in unstressed cells, which is believed to include a role as a “molecular chaperone.” The hair cycle is characterized by rhythmic tissue remodeling processes, and is an intriguing model for studying the relation between keratinocyte differentiation and HSP expression under physiologic circumstances. We have therefore studied, by immunofluorescence, the expression of selected HSPs during the murine hair cycle. Methods The association between hair follicle cycling and the expression of three selected HSPs (HSPs 27, 60, and 72) was examined by immunofluorescence, using the depilation-induced hair cycle of C57BL76 mice. Results HSP expression was absent from telogen follicles, and was restricted predominantly to keratinocytes in the bulge and the cycling epithelial portion of the hair follicle during anagen and catagen. Immunoreactivity for HSPs 27, 60, and 72 in the hair bulb increased significantly during anagen VI and the catagen transformation of the follicle, and decreased again abruptly with completion of the catagen-telogen transformation. The expression pattern of HSPs 60 and 72 in situ was cytoplasmic, whereas that of HSP 27 was both cytoplasmic and nuclear. Conclusions These observations suggest that the synthesis of HSPs by hair bulb keratinocytes is related to the anagen-catagen transformation of the follicle, possibly reflecting keratinocyte apoptosis and/or terminal differentiation in the regressing, cycling portion of the follicle. In addition, the rather proximal localization of HSP expression makes it unlikely that the HSPs examined interact with the more distally located intrafollicular γ/δ T-cell receptor-positive lymphocytes.